Tumour-induced osteomalacia (TIO) is a rare paraneoplastic syndrome characterised by severe hypophosphataemia and osteomalacia, with renal phosphate wasting that occurs in association with tumour. The epidemiology likewise aetiology is not known. The clinical presentation of TIO includes bone fractures, bone and muscular pains, and sometimes height and weight loss. TIO may be associated with mesenchymal tumours which may be benign or malignant in rare cases. Mesenchymal tumour itself may be related to fibroblast growth factor 23 (FGF23), which is responsible for hypophosphataemia and phosphaturia occurring in this paraneoplastic syndrome. Hypophosphataemia, phosphaturia and elevated alkaline phosphatase are the main laboratory readings that may lead to more precise investigations and better diagnosis. Finding the tumour can be a major diagnostic challenge and may involve total body magnetic resonance imaging, computed tomography and scintigraphy using radiolabelled somatostatin analogue. The treatment of choice for TIO is resection of a tumour with a wide margin to insure complete tumour removal, as recurrences of these tumours have been reported. We provide here an overview on the current available TIO case reports and review the best practices that may lead to earlier recognition of TIO and the subsequent treatment thereof, even though biochemical background and the long-term prognosis of the disease are not well understood. This review also includes a 4-year-long history of a patient that featured muscular pains, weakness and multiple stress fractures localised in the hips and vertebra with subsequent recovery after tumour resection. Because the occurrence of such a condition is rare, it may take years to correctly diagnose the disease, as is reported in this case report. Tumour-induced osteomalacia (TIO) is a rare paraneoplastic syndrome clinical presentation of which includes bone fractures, bone and muscular pains, and sometimes loss of height and even weight. Weight loss is unusual, but sometimes observed, and could be explained by general debilitated state of the patient with consequent poor nutrient intake and loss of muscle mass , 2. MostTIO is characterised by severe hypophosphataemia and osteomalacia, with renal phosphate wasting that occurs in association with tumour. TIO also is called oncogenic hypophosphatemic osteomalacia (M83.8) and appears in the portal for rare diseases and orphan drugs under that name (ORPHA352540). It was first described in 1947 by Robert McCance, who reported a patient with pain, weakness, gait abnormalities, and low phosphorus levels. The patient was treated with high doses of vitamin D, but the symptoms did not completely resolve until a tumour in femur bone was removed [The first person to clearly recognise that the disease was the result of a “rachitogenic” substance was Andrea Prader. In 1959, he described an 11 1/2-year-old girl who developed severe rickets over the course of a year. Evaluation showed decreased tubular phosphate reabsorption but otherwise normal kidney function. A tumour, classified as a giant cell granuloma, was identified in a rib and removed resulting in healing of rickets .The prevalence of the disease is not known. Since the association between phosphate reabsorption and tumour was first made, more than 300 cases of TIO have been reported in the literature .The first evidence of a circulating factor that could cause phosphate wasting in phosphaturic disorders such as TIO was demonstrated in a set of experiments in mice by Meyer et al. and Nesbitt et al. , 18. TheFGF23 was first identified as the phosphaturic substance when mutations in FGF23 were linked to autosomal-dominant hypophosphatemic rickets (ADHR) , 21. SooExcessive FGF23 action causes several hypophosphatemic diseases, including TIO, X-linked hypophosphatemic rickets (XLHR), autosomal dominant and recessive hypophosphatemic rickets (ADHR and ARHR) –28. UndeThe tumours associated with TIO are usually small and mesenchymal in origin . The proWeidner in 1991 was the first to propose a classification system based on the histological findings of 16 cases of TIO and designated the tumours as phosphaturic mesenchymal tumours . TIO tumWhen testing antigen expression, FGF23 is positive in about 70 % of all the cases studied, and the proliferating cells within the tumour are usually the source of FGF23 –35.While typically benign, malignant presentation and metastases can occur , 5, 36. Regardless of tumour morphology, the hallmark of the diagnosis is the association of the tumour with the clinical syndrome of TIO, which includes an elevation in plasma FGF23 and its disappearance after tumour resection.Quoting Jiang et al. who reviewed 308 tumour-induced osteomalacia cases reported in English literature between 1987 and 2011, about 46 % of reported cases of TIO have occurred in females and 56 % in males, with a mean age of 45.3 years when definitive diagnosis was made . Within Patients with TIO often present with many years of symptoms before they are diagnosed. The symptoms usually are nonspecific and often progressive. Common complaints are bone pain, muscle weakness, reduced height, and multiple fractures, primarily in the ribs, vertebral bodies, and femoral neck , 37. TheDifferential diagnosis should always include renal Fanconi’s syndrome—a disorder of the proximal renal tubules, leading to impaired phosphate reabsorption and hypophosphatemia. This syndrome may be genetic in origin or it may occur as a complication of myeloma, amyloidosis, or Sjogren’s syndrome. It can also occur with certain medications or heavy metal poisoning. The diagnosis can be confirmed by normal levels of FGF23 along with the presence of glycosuria, hypokalaemia, and metabolic acidosis [Once the diagnosis of an FGF23-dependent, phosphate wasting disorder is made, a thorough history can aid in excluding the genetic causes, such as XLHR, ADHR and ARHR. Genetic testing can also be done.Having narrowed the diagnosis to TIO, a careful physical examination should be performed, as the tumours that cause TIO can sometimes be found in the subcutaneous tissue . As tumoFinding the tumours can be a major diagnostic challenge and may involve total body magnetic resonance imaging (MRI); as tumours can occur anywhere in the body, it is important to scan the whole body, including extremities, which is often excluded in routine nuclear medicine imaging , compute111In-labelled pentatreotide. Octreotide scintigraphy is successfully used to locate tumours in up to 95 % of patients with TIO [A stepwise approach is advocated, first performing functional tests. Tumours associated with osteomalacia variably express five somatostatin receptors (SSTR1-5), allowing SSTR-based functional imaging by somatostatin analogue scintigraphy or positron emission tomography , 52, 53.with TIO . It may with TIO . Despitewith TIO . Somatoswith TIO .Anatomic imaging  should be performed to confirm the location of the tumour after suspicious lesions had been identified by functional imaging . High-reDespite all of the advances in imaging that are available today, tumour localization may not be successful. If this is the case, imaging studies should be repeated every 1–2 years.The treatment of choice for TIO is resection of a tumour with a wide margin to insure complete tumour removal, as recurrences of these tumours have been reported , 9, 36. In case of incompletely resected tumours, subsequent radiotherapy can be used to avoid recurrence or metastasis (41). Late recurrence due to metastatic disease is rare but possible. This probably occurs in less than 5 % of the patients with TIO , 36. LunWhen the tumour cannot be localised or is not surgically resectable, medical therapy with phosphate supplementation and calcitriol or alfacalcidol is used. Treatment with octreotide is an alternative form of medical therapy that may be considered. In one case report, phosphate wasting and hypophosphatemia were corrected with octreotide therapy; this effect was presumably mediated by somatostatin receptor expression by the tumour, which was demonstrated by octreotide scintigraphy . HoweverAnti-FGF23 antibody is being studied as a novel therapy for FGF23-related hypophosphatemic diseases . ResultsThe prognosis depends on detecting the tumour and possibility to remove it widely. The tumours when detected are typically benign in most of the cases. The symptoms disappear and the healing of the bones begin after total removal of the tumour. Nevertheless, the follow-up should be continued because the delayed metastasis can occur as it was described in few cases.2). Physical examination revealed scoliosis and kyphosis of the spine, overall weakness, weakness in muscles and pain in both arms when palpating. The laboratory investigations showed no inflammation, and the only abnormality in serological readings was elevated alkaline phosphatase—248 U/l  due to its bone-specific fraction comprising 80.5 %  and weighted 73 kg  was documented for the first time. The 24-h urine sample phosphaturia showed normal phosphate clearance 26.5 mmol/24 h  and subsequent phosphate readings in urine were within normal values as well. Vitamin D levels, calcium and ionised calcium readings were found within the normal values: vitamin D being 78.4 nmol/l ; calcium 2.25 mmol/l ; and ionised calcium 1.12 mmol/l ; nevertheless, he used periodically prescribed bone mineral supplements from the beginning of disease. The radiology of spine showed a new compressive fracture in the level Th7–Th8 in addition to the old fracture, which was later confirmed by CT scan. Whole body bone scintigraphy with 99mTc-MDP 550 MBq showed increased focal uptake of a radiotracer in the left shoulder and in the ribs on both sides due to osteoblastic process and reduced uptake in right femoral on SPEC/CT sclerotic bone lesion without tumour at that time or Morbus Paget specificity  administered intravenously and repeated after a year of interval but with no clinical or laboratory improvement. It was discontinued and switched to oral bisphosphonate. Natrium risedronate was initially prescribed in dosage of 35 mg once per week and subsequently increased to 35 mg per day but was discontinued in the fall 2014 because of no effect.In February 2012, the avascular necrosis of both femur heads with osteosclerotic locus in the right femur of unknown origin was found by MRI. During routine follow-up in 2013, additional compressions were found in spine CT, now overtaking the segment from T5 to T12. The bone density continued to diminish up to 0.76 g/cm2), permanent pain in chest and hips, progressing breathing discomfort, muscular weakness, inability to bend below knees and posture change , lead 9.60 μg/l , cadmium 0.8 μg/l  and mercury <1.0 μg/l . Because of disease manifestation in adulthood, no analysis for FGF gene mutation was requested. FGF-23 readings were found increased fivefold and being 589 U/l . The whole body scintigraphy with somatostatin analogues 99mTc-tekrotyd 605 MBq revealed intensive uptake of radiotracer in the right femoral head, and SPECT/CT demonstrated uptake in sclerotic bone lesion in the right femoral head—tumour intensively expressing somatostatin receptors . The alkaline phosphatase returned to normal values 6 months after tumour extirpation and remained within normal values after 1 year of follow-up  with bone fraction now standing at 57.7 % and FGF-23 returned to normal values 104 IU/l (26–110 IU/l). Three months after surgical procedure, the body pain almost disappeared and the weight increased up to 62 kg and height up to 166 cm (BMI = 22.5 kg/mThe diagnosis of TIO is a challenge under the best of circumstances. TIO should be included in the differential diagnosis in patients with progressive weakness, bone and muscle pain and multiple fractures. The diagnosis is commonly delayed for years due to the nonspecific nature of the presenting symptoms, failure to include determination of serum phosphorus levels in blood chemistry testing and difficulty in identifying the responsible tumour. The professional evaluation of whole body images, including single-photon emission tomography enables identification and better tumour contrast which can be found in skeletal structures of whatever region of the human body. FGF23 may be of help to understand the underlying mechanisms of TIO and a measurement to follow up the disease course.This review includes the description of one case report, and the consent form for publication was obtained from the person. The consent form of that particular person was sent to the Editor of this journal.


Adropin is a 4.9 kDa peptide that is important for maintenance of metabolic and non-metabolic homeostasis. It regulates glucose and fatty acid metabolism and is involved in endothelial cell function and endothelial nitric oxide (NO) synthase bioactivity as well as physical activity and motor coordination. Adropin is expressed in many tissues and organs including central nervous system (CNS). This peptide plays a crucial role in the development of various CNS disorders such as stroke, schizophrenia, bipolar disorder as well as Alzheimer's, Parkinson's, and Huntington's diseases. In this comprehensive review, the potential roles of adropin in cellular signaling pathways that lead to pathogenesis and/or treatment of CNS disorders will be discussed. Enho) located on chromosome 9 —a rate-limiting enzyme in glucose oxidation. Further, adropin suppresses two key enzymes in fatty acid utilization: carnitine palmitoyltransferase-1B (CPT-1B) and Cluster of Differentiation 36 (CD36)  Figure . In thisVEGFR2—a tyrosine kinase receptor—is especially expressed in endothelial cells and regulates endothelial function and angiogenesis. Adropin strongly upregulates this receptor, activates PI3K/Akt and ERK1/2 pathways, and enhances eNOS thus, modulating NO bioavailability  under the effect of growth factors such as VEGF, cytokines, insulin, and other cellular stimuli Figure . ActivatPhosphorylated-Akt provokes cell cycle progression, proliferation, differentiation, and survival  and haplotype studies indicated the involvement of Akt in Schizophrenia  and reduction in anti-apoptotic factors (BDNF and Bcl-2) and the synaptic markers (synaptophysin and drebrin)  Figure . BNDF prOne of the endothelial protective functions of adropin is regulation of nitric oxide (NO) bioavailability  that is upregulated by PI3K/Akt and ERK 1/2 signaling pathways Figure . NOS polin-vivo and in-vitro endothelial cells resulting in proliferation, migration, and capillary-like tube formation and diminished permeability and apoptosis of these cells  in the vicinity of plaques of Alzheimer's disease, and in the cerebrovascular wall of hemorrhagic stroke had been described  and down-regulating pyruvate dehydrogenase kinase-4 (PDK-4)- a PDHC inhibitor. PDHC is a mitochondrial matrix enzyme complex that catalyzes oxidative decarboxylation of pyruvate to produce acetyl-CoA, which plays a critical role in cerebral aerobic energy metabolism (Cardell et al., The involvement of Adropin in endothelial permeability was originally described by Lovren and coworkers in 2010 (Lovren et al., Induction of mesenchymal cells with inhibitors of prolyl hydroxylase—a key enzyme in HIF-1α degradation—promotes mesenchymal cells differentiation to morphologically neuron-like cells (Pacary et al., Recent studies demonstrated that inhibition of Rho/ROCK signaling pathway enhances HIF-1 activity and upregulates EPO, VEGF and p21, and consequently potentiates neurogenesis (Pacary et al., Stein et al. discovered GPR19 as a potential adropin receptor (Stein et al., Studies regarding the effects of adropin in different organs are still in infancy stage, but increasing evidence suggest that this peptide has unique effects on endothelial cell function via upregulating eNOS expression through the VEGFR2-PI3K-Akt, VEGFR2-ERK 1/2 pathways and inhibition of Rho/ROCK pathway. However, our current knowledge mainly comes from animal studies or treatment with the putative secreted domain of adropin. Whether these findings are transferable to clinical studies needs to be determined. Moreover, adropin may be utilized as a promising biomarker for CNS disease risk stratification or diagnosis, and/or a potential therapeutic candidate in CNS injuries. Although adropin seems to be a novel target to limit vascular diseases, in parallel with the documented effects on metabolic modulation, further investigations are needed to elucidate the specific mechanism underlying the association between adropin and CNS diseases.Study concept and design: SA, RZ. Acquisition of data: SS, SA, TP, RZ. Drafting and critical revision of manuscript: SS, SA, TP, RZ. Study supervision: TP, RZ.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Clostridium infiltrate and selectively germinate in these necrotic regions, providing cancer-specific colonisation. The specificity of this system was first demonstrated over 60 years ago and evidence of colonisation has been confirmed in multiple tumour models. The use of “armed” clostridia, such as in Clostridium Directed Enzyme Prodrug Therapy (CDEPT), may help to overcome some of the described deficiencies of using wild-type clostridia for treatment of cancer, such as tumour regrowth from a well-vascularised outer rim of viable cells. Successful preclinical evaluation of a transferable gene that metabolises both clinical stage positron emission tomography (PET) imaging agents  as well as chemotherapy prodrugs  would be a valuable early step towards the prospect of “armed” clostridia entering clinical evaluation. The ability to target the immunosuppressive hypoxic tumour microenvironment using CDEPT may offer potential for synergy with recently developed immunotherapy strategies. Ultimately, clostridia may be most efficacious when combined with conventional therapies, such as radiotherapy, that sterilise viable aerobic tumour cells.Most solid cancers contain regions of necrotic tissue. The extent of necrosis is associated with poor survival, most likely because it reflects aggressive tumour outgrowth and inflammation. Intravenously injected spores of anaerobic bacteria from the genus  Solid tumours account for approximately 90% of all diagnosed cancer , and theClostridium is one of the largest prokaryotic genera, comprising a heterogeneous group of rod-shaped, anaerobic, spore-forming bacteria. In times of stress, they can undergo a complex cell differentiation process resulting in the production of endospores, rendering them highly resistant to harsh environmental conditions such as high temperature and dehydration -fluoro-3-[hydroxymethyl]butyl)-guanine ([18F]-FHBG) is an example of a metabolite that is trapped inside transduced cells and detectable as early as two hours after administration [18F]-FHBG as the first investigational new imaging reporter probe by the U.S. Food and Drug Administration (FDA) [18F]-FHBG in tumour nodules treated by direct injection of an adenoviral vector encoding HSV-tk [18F]-FHBG and concentrate the radiotracer before the cell is lysed by the virus. In principle, this would not be a problem for clostridia, which germinate autonomously in the necrotic areas of tumours.Although no work has yet been published describing PET reporter systems specifically for imaging  by hNIS . A high  by hNIS , and non by hNIS ,53. Anot by hNIS . 9-(4-[1stration . Preclinon (FDA) , and stug HSV-tk . HoweverPET probes able to penetrate into human tumours and detect a distant subset of hypoxic cells clearly have adequate pharmacokinetic and pharmacodynamic properties for molecular imaging, and, as such, the probes have attained a high level of validation ,58,59. IActivation of the immune system as a treatment modality for cancer has long been a goal in oncology. Although insight into anti-tumour immune responses has significantly increased in recent times, development of immunotherapy for therapeutic benefit in cancer treatment has faced several difficulties, including defining the optimal dose/schedule/patient population and inadequate methods for evaluating efficacy. However, recent proof of clinical efficacy studies in patients treated with several immunotherapy drugs has increased clinical development of cancer immunotherapy ,62.C. novyi-NT is immunogenic in syngeneic mouse CT26 colorectal and rabbit VX2 hepatoma tumour models, eliciting a strong inflammatory response with influx of neutrophils and other inflammatory cells [C. novyi-NT cured subjects, suggesting the innate immune response eventually evolves into an adaptive immune response. These observations suggest a germination-dependent stimulation of an immune response with pathogen-associated antigen recognition acting as an adjuvant.Clostridial endospores are themselves poorly immunogenic and fail to elicit an immune response, persisting in multiple organs following intravenous injection . In contry cells . LocalisC. acetobutylicum has previously been engineered to express increased levels of IL-2 and sufficient levels of IL-2 were generated to produce an anti-tumour effect whilst avoiding the side effects demonstrated using systemic administration [C. novyi-NT variant in tumour tissue [Clostridia also have the potential to deliver immunotherapy agents directly to tumour tissue . For exastration ,70. In ar tissue . Thus, tCDEPT may also be combined with cancer immunotherapy. Here, local redistribution of cytotoxic prodrug metabolites results in cell kill in the perinecrotic hypoxic regions, generating local tumour antigen release. Moreover, the hypoxic microenvironment plays an important role in compromising immune cell function, driving immune suppression, and T cell exclusion . TargetiC. novyi-NT confirms the propensity for human tumour colonisation. We propose that non-invasive imaging is fundamental to the successful clinical development of live biotherapeutic agents, as it provides vital real-time biomarker tracking in early safety trials and rational expansion of subsequent efficacy studies. Development of a clostridial vector with PET-imaging capacity could provide a platform technology for the delivery of many biological agents, thereby turning a pathological feature associated with treatment failure into a precision therapy.Tumour necrosis is most commonly seen in advanced and end-stage cancers where few, if any, treatment options remain. Thus, armed clostridial vectors may represent a valuable salvage therapy when other treatment options are exhausted. Historical clinical evaluation supports proof of concept and recent progress with


Lubrication plays an important role in the clinical performance of the ceramic-on-ceramic (CoC) hip implant in terms of reducing wear and avoiding squeaking. All the previous lubrication analyses of CoC hip implants assumed that synovial fluid was sufficiently supplied to the contact area. The aim of this study was to investigate the lubrication performance of the CoC hip implant under starved conditions. A starved lubrication model was presented for the CoC hip implant. The model was solved using multi-grid techniques. Results showed that the fluid film thickness of the CoC hip implant was affected by fluid supply conditions: with the increase in the supplied fluid layer, the lubrication film thickness approached to that of the fully blooded solution; when the available fluid layer reduced to some level, the fluid film thickness considerably decreased with the supplying condition. The above finding provides new insights into the lubrication performance of hip implants. The ceramic-on-ceramic (CoC) hip implant is increasingly used due to its outstanding tribological and biocompatible properties. Lubrication plays an important role in the clinical performance of the CoC hip implant. First, poor lubrication is one potential reason of squeaking noise, which is an audible phenomenon receiving increasing concerns , of CoC However, all the previous lubrication analyses of CoC hip bearings were based on an assumption that synovial fluid is sufficiently supplied to the lubricated contact area . This is22.1A typical CoC total hip replacement bearing, which consists of three components, a titanium acetabular shell, a ceramic insert and a ceramic head, was considered. The ceramic insert is normally fixed in the titanium acetabular shell using a taper locking mechanism. The initial stability of the acetabular shell is achieved using either cemented or uncemented methods while the long term fixation is reached by the in-growth of bone onto and around the porous-coated shell surface. The spherical ceramic head articulates against the hemi-spherical inner surface of the ceramic insert to form a joint. In the present study, the insert was assumed to be securely fixed to the shell. A uniform thickness of 10 mm and 4 mm was adopted for the ceramic insert and the titanium shell, respectively. The bone and the fixation of the shell were represented by an equivalent support layer with a thickness of 2 mm and appropriate material properties . Such a 5/s), it was reasonable to assume the synovial fluid as Newtonian, isoviscous and incompressible  and the gap height (h) , θf=1.The governing equations of the lubrication model were established in spherical coordinates . The stardinates , and θf ight (h) :(2)θf or in a starved region (p=0 and θf<1).Besides the boundary conditions similar to the fully flooded lubrication :(3)pp; ex and ey are eccentricities of head relative to the cup; δ is the elastic deformation of the bearing surfaces, determined by the deformation coefficients of the bearing surfaces and the film pressure.The total gap equation consisted of the undeformed gap and the elastic deformation of bearing surfaces due to the film pressure:In addition, the external load components were balanced by the integration of the film pressure:2.3θ and ϕ directions  and starved (p=0) regions can be found in θf=1; while in the starved region, 0<θf<1. In addition, the fractional film content had a jump at the inlet meniscus, which was at the same location where the fluid started pressurizing (p>0 and θf=1), the fluid film thickness was the same as the total gap, while out of the pressurized region (p=0 and 0<θf<1), the fluid film thickness was less than the total gap. This was consistent with the definition of θf. The above consistencies suggest that the model and solution in the present study are reasonable.The typical three-dimensional distributions of the fluid pressure, fractional film content and fluid film thickness of the CoC hip implant under starved conditions are shown in surizing . Correspsurizing . These osurizing  showed thfluid=0.2 µm, hfluid=2.0 µm and the fully flooded condition were 0.072, 0.073 and 0.075 µm, respectively (hfluid=0.14 µm in this study), the fluid film thickness considerably decreased with the fluid supply condition , and geometric parameters should be analyzed in the future.Lubrication analysis has played an important role in the development of hip implants through investigating how the geometric parameters and working conditions affect the fluid film thickness . HoweverThere are, of course, limitations in the present study as discussed above. For example, as the first step towards investigating the starved lubrication of CoC hip implants, only the steady state condition and one group of loading and geometric parameters were considered. Moreover, to achieve a good convergence, a smaller radial clearance was used. Despite the above limitations, the present study has paved the way towards fully understanding lubrication performance of the CoC hip implant under starved lubrication and provided important initial understanding.5A starved lubrication model was presented and solved for the CoC hip implant for the first time. Results showed that for the conditions considered in this study, the lubrication performance of the CoC hip implant was affected by fluid supply: with the increase in the supplied fluid layer, the lubrication film thickness approached to that of the fully blooded solution; when the available fluid layer reduced to some level, the fluid film thickness considerably decreased with the reduction of fluid supplied to the inlet. Such a variation in the fluid film thickness with starved conditions implies the importance of considering starved lubrication in the CoC hip implant.


Twitter is a popular microblogging platform for the rapid dissemination of information and reciprocal exchange in the urological field. We aimed to assess the activity, users and content of the online discussion, #KidneyStones, on Twitter.We investigated the Symplur Signals analytics tool for Twitter data distributed via the #KidneyStones hashtag over a one year period. Activity analysis reflected overall activity and tweet enhancements. We assessed users’ geolocations and performed an influencer analysis. Content analysis included the most frequently used words, tweet sentiment and shares for top tweets.3,426 users generated over 10,333 tweets, which were frequently accompanied by links (49%), mentions (30%) and photos (13%). Users came from 106 countries across the globe and were most frequently from North America (63%) and Europe (16%). Individual and organisational healthcare professionals made up 56% of the influencers of the Twitter discussion on #KidneyStones. Besides the words ‘kidney’  and ‘stones’ , ‘pain’ , ‘urine’ , and ‘risk’  were the most frequently used words. 56% of tweets had a positive sentiment. The median (range) number of shares was 85 (62–587) for the top 10 links, 45.5 (17–94) for the top 10 photos, and 44 (22–95) for the top 10 retweets.The rapidly growing Twitter discussion on #KidneyStones engaged multiple stakeholders in the healthcare sector on a global scale and reached both professionals and laypeople. When used effectively and responsibly, the Twitter platform could improve prevention and medical care of kidney stone patients. The microblogging social media platform, Twitter, enjoys increasing popularity in the healthcare sector. Currently, over 70% of urologists in Australia and New Zealand have a social media presence, with Twitter being the second most commonly used form after LinkedIn, which serves a completely different purpose . UrologiIn slightly more than a decade, the prevalence of urolithiasis in the United States has increased from 5.2% to 8.8%  and incrIn the current investigation, we aimed to assess the potential of Twitter to constitute a potential platform for the dissemination of contemporary evidence on prevention, diagnosis and treatment of stone disease. Hypothesising that a Twitter discussion on #KidneyStones might appreciably involve both healthcare givers and laypeople, as well as encourage considerable global public awareness, we investigated the characteristics of its activity, users and content.www.symplur.com) is a Twitter analysis website that maintains a database of healthcare-related Twitter conversations. Symplur Signals  is a fee-based research analytics tool that promotes the understanding of healthcare as seen by patients, doctors and other stakeholders with access to healthcare social media data points.The study was approved by the Ethics Commission of the Faculty of Medicine of the University of Cologne. We performed an extensive analysis of activity, users and content of the online discussion on #KidneyStones on Twitter, using the Symplur Signals database. Symplur  to assess the issues inducing peak activities. The tweet metrics analysis was performed by retrieving statistics about ratio and frequency of retweets, tweets with links, tweets with photos, tweet replies and tweets where one or more Twitter users were mentioned. Engagement metrics were retrieved by obtaining the number of users who tweeted over a set period of time, grouped by the number of tweets sent. A tweet language analysis illustrated the language used by active participants over a set period of time. Language type was identified by a natural language processing algorithm directly provided by the Twitter application programming interface.User analysis included the cumulative user report, users’ geolocations, and an influencer analysis of the top influencers in the #KidneyStones discussion. Cumulative and new users were recorded for monthly intervals. Cumulative users did not represent recurring users, but were counted as a new user the first time they used the #KidneyStones hashtag and as a previous user in subsequent reporting periods, regardless of activity. The geolocation of users was recorded when users sent tweets tagged with certain geolocation data. We analysed the top 100 contributors to the #KidneyStones discussion, as measured by number of tweets in the influencer analysis. For this purpose, we performed a Twitter profile analysis and assigned these top influencers to these healthcare categories, in line with the Symplur Signals healthcare category definitions: physician; patient; healthcare professional; caregiver/advocate; researcher/academic; individual other healthcare; individual other non-healthcare; organisation provider; organisation research/academic; organisation government; organisation advocate/support; organisation pharma; organisation other healthcare; organisation other non-healthcare, and spam .We used the Symplur Signals tools for content analysis. The 100 most frequently used words in tweets on #KidneyStones were analysed and counted. Since multiple hashtags are often used within a single tweet, we used the hashtag network graph to analyse hashtags accompanying the #KidneyStones discussion and their relationships. The sentiment report analysed tweets for positive and negative sentiment by a natural language processing algorithm. The algorithm is based on two custom dictionaries, one for positive words and one for negative words. Each word in the dictionaries has a weighting from one to five, with five being the highest. Finally, we investigated the most frequently shared links, photos and the most frequently retweeted tweets. We performed statistical calculations using the Statistical Package for the Social Sciences 22.0 software . Values are described as median and range.Out of 10,333 total tweets, 5,013 (49%) were accompanied by links, 3,101 (30%) by mentions and 1,337 (13%) by photos. 2,441 (24%) were retweets and 229 (2%) were tweet replies. 2,798 users (82%) generated one tweet, 331 (10%) two, 244 (7%) three to nine, and 53 (2%) 10 or more tweets. 9,942 (96%) of tweets were in the English language.A content analysis of the most frequently used words in the #KidneyStones online discussion on Twitter is shown in We performed a longitudinal assessment of activity, users and content of the online discussion on #KidneyStones, using the Symplur Signals health analytics database. Over the one year period, Twitter activity was reflected by 10,806 tweets, which were frequently enhanced with links and mentions, and were posted predominantly in English. Together, 3,426 users from 106 countries contributed to the online discussion, with non-professional individuals and healthcare organisations being the main influencers. Content was dominated by the words ‘kidney’, ‘stones’, ‘pain’, ‘urine’ and ‘risk’, and tweets had more positive than negative sentiments. The top links, photos and retweets were shared up to 100 times.The Twitter activity of 10,806 tweets found for #KidneyStones in the study period is much higher than 880 tweets found in 2012 for urinary tract infection, another urological disease . The majThe tweet metric analysis showed that contributors interacted using retweets (24%) and mentions (30%) in their tweets. Relevant tweets inducing retweets are of high interest to the Twitter community, which is underlined by the recently introduced concept of the Twitter impact factor . MoreoveThe Twitter discussion on #KidneyStones is global, involving users from all continents. Usage rates of social media were 74% among American urologists in 2013  and 70% Particularly noteworthy is the finding of the content analysis, revealing that words symbolising disease awareness and prevention are most frequently used in tweets on #KidneyStones. Primary prevention of kidney stones has been shown to be cost-effective for a national healthcare system . This isAlthough the #KidneyStones hashtag is proposed as standardised communication descriptor , we acknWe also used the Symplur Signals analytics platform for a systematic assessment of healthcare social media data. The automated data extraction and analysis algorithms allow for the analysis of a vast amount of data, but cannot detect linguistic nuances, such as ambiguity or irony when analysing content. Lastly, Twitter is a rapidly growing and changing social media platform, implying that the results of our contemporary analysis might be out-dated in the near future.Notwithstanding the aforementioned limitations, several practice-oriented conclusions can be drawn from the findings of the current study. The Twitter discussion on #KidneyStones is maintained by users from all over the world and evokes a remarkable number of tweets, underscoring the global reach of this microblogging platform. Healthcare organisations, as one of the top influencers in the discussion, have a unique opportunity to raise the awareness of patients and providers for nonmedical and pharmacological prevention, eventually reducing recurrence rates and care-related expenditure. Patients and other laypeople substantially contributing to the discussion have the option of being discreetly and noncommittally counselled by experts, optimising shared decision-making. Finally, given responsible Twitter usage, the dissemination of novel diagnostic and therapeutic developments in the area of urolithiasis between stakeholders and patients might be considerably accelerated.The Twitter discussion on #KidneyStones engaged multiple stakeholders in the healthcare sector on a global scale and involves both professionals and laypeople. Considering the rapidly increasing prevalence and treatment-related costs of urolithiasis, Twitter might promote shared decision-making and contribute to the optimisation of patient care.S1 Table(XLSX)Click here for additional data file.S2 Table(XLSX)Click here for additional data file.


The diagnosis of brain glioma can challenging, particularly when the masses are very large. While the majority of gliomas are astrocytomas or oligodendrogliomas, enough are of other types that awareness of these others is critical. This course will begin with a review of some of the distinctive features of common and less common brain tumours, with particular focus on the distinctions that make a difference in approach.Once a tumour is diagnosed and therapy instituted, clinicians will appreciate it greatly if you interpret the images with a mind to how they measure therapy response. This requires an awareness of the treatment regimen the patient is on, and changes in agents like steroid dose.RECIST is applied throughout most of the body, but recently, the Response Assessment in NeuroOncology (RANO) criteria were published, to address some of the specifics of neurooncology . These aRANO is largely focused on conventional anatomic imaging methods, but new quantitative methods reflecting water diffusion and tumour perfusion appear to improve response assessment. Diffusion restriction typically reflects higher cell density seen in viable tumour as well as higher grade tumour . ChangesThe combination of temozolomide and radiation has been shown to improve survival but also has a high rate of pseudoprogression, which is present in 1/3 to 1/2 of subjects, particularly when the tumour is MGMT methylated . It is i


Perioperative fluid therapy remains a highly debated topic. Its purpose is to maintain or restore effective circulating blood volume during the immediate perioperative period. Maintaining effective circulating blood volume and pressure are key components of assuring adequate organ perfusion while avoiding the risks associated with either organ hypo- or hyperperfusion. Relative to perioperative fluid therapy, three inescapable conclusions exist: overhydration is bad, underhydration is bad, and what we assume about the fluid status of our patients may be incorrect. There is wide variability of practice, both between individuals and institutions. The aims of this paper are to clearly define the risks and benefits of fluid choices within the perioperative space, to describe current evidence-based methodologies for their administration, and ultimately to reduce the variability with which perioperative fluids are administered.Based on the abovementioned acknowledgements, a group of 72 researchers, well known within the field of fluid resuscitation, were invited, via email, to attend a meeting that was held in Chicago in 2011 to discuss perioperative fluid therapy. From the 72 invitees, 14 researchers representing 7 countries attended, and thus, the international Fluid Optimization Group (FOG) came into existence. These researches, working collaboratively, have reviewed the data from 162 different fluid resuscitation papers including both operative and intensive care unit populations. This manuscript is the result of 3 years of evidence-based, discussions, analysis, and synthesis of the currently known risks and benefits of individual fluids and the best methods for administering them.The results of this review paper provide an overview of the components of an effective perioperative fluid administration plan and address both the physiologic principles and outcomes of fluid administration.We recommend that both perioperative fluid choice and therapy be individualized. Patients should receive fluid therapy guided by predefined physiologic targets. Specifically, fluids should be administered when patients require augmentation of their perfusion and are also volume responsive. This paper provides a general approach to fluid therapy and practical recommendations. Major surgery is a considerable physiologic insult that can be associated with significant morbidity and mortality. The occurrence of one or more postoperative complications adversely effects both short-term and long-term survival and increases healthcare costs ,2. The pPerioperative fluid management remains a highly debated topic. There is wide variability of practice, both between individuals and institutions. Perioperative morbidity is linked to the amount of intravenous fluid administered (fluid therapy) with both insufficient and, more commonly, excess fluid delivery leading to increased postoperative complications -5. CurreMultiple studies have shown that approaching fluid therapy with the goal of hemodynamic stabilization can reduce complications after major surgery -9. More A patient’s physiologic status in general and hemodynamic stability in particular define the need for cardiovascular support, including fluid therapy and use of vasoactive drugs  and inotropes. Specific hemodynamic goals include maintaining adequate blood volume and sustaining perfusion pressure so as to maintain cardiac output, tissue blood flow, and adequate oxygen delivery. Fluid therapy is often the first line of hemodynamic support because decreased effective circulating blood volume often accompanies induction of anesthesia and surgical trauma. However, fluid therapy only indirectly impacts cardiac and vascular function. Optimizing oxygen delivery and assuring the removal of metabolic bioproducts may require a combination of individualized fluid therapy, pharmacotherapy, and occasionally mechanical cardiovascular support.Fluid infusions directly increase vascular volume, subsequently and usually improve global and regional perfusion and blood pressures if the heart is preload-responsive, and often improve oxygen delivery and tissue oxygenation. However, these changes are profoundly influenced by the cardiac and peripheral vascular status . Thus, tIntravenous infusion of fluid directly expands plasma volume with transient or sustained effect that varies based on the colloid osmotic properties of the fluid, blood flow distribution, type and level of anesthesia, vascular endothelial integrity, and the physiologic state. The expansion of plasma volume causes the mean systemic pressure to increase, and, if greater than right atrial pressure, the pressure gradient for venous return will increase. If the right and left ventricle are volume responsive, then cardiac output will also increase. There is no easy means to measure plasma volume nor is there a defined means of how measured plasma volume could be used to achieve the physiologic goals of optimal pressure, flow, and oxygenation for the perioperative patient. Although one can estimate mean systemic filling pressure in appropriately instrumented patients, it is unclear if such measures will alter either therapy or outcome, because knowing plasma volume and even effective circulating blood volume only gives a partial picture of the determinants of cardiac output. Other critical factors include blood flow distribution, vasomotor tone, right ventricular function, and the level of positive-end expiratory pressure, which, individually and collectively, may alter cardiovascular responsiveness. Given the absence of easily obtainable regional measures of perfusion, the anesthesiologist may consider assessing global perfusion by measuring base deficit, lactate, and central and mixed venous oxygen saturation to clarify the impact of selected interventions.Perioperative assessment of changes in blood volume is difficult and requires evaluation of several clinical and physiologic events that accompany major surgery. Standard hemodynamic monitoring devices fail to detect occult hypovolemia , which oTachycardia is considered a classic sign of hypovolemia, but the assessment of intravascular volume based on heart rate lacks sensitivity and specificity  for a vaPerioperative hypovolemia is deleterious to organ function because normal adaptive mechanisms cause peripheral vasoconstriction to sustain blood flow to the heart and brain, causing ischemia to other organ and surgical tissues in need of blood flow for repair. For surgical patients, several factors including preoperative fasting, hypertonic bowel preparations, anesthetic agents, and positive pressure ventilation all contribute to reduced effective circulating blood volume. Anesthetized patients often present with a functional intravascular volume deficit . On the a priori perioperative fluid plan. Taken as a whole, the success of both GDT and of some restrictive fluid strategies suggests that perioperative fluid planning must emphasize that fluid therapy be administered only with clear indication. Functional hemodynamic parameters offer unique information about fluid responsiveness, which my help detect fluid needs and avoid unnecessary fluid loading. Despite their limitations and confounding factors, this information may be crucial in guiding fluid therapy in surgical patients [Establishing what constitutes a restrictive or liberal amount of fluid from the literature is difficult because the absolute amounts of fluid administered vary substantially among trials making any conclusion difficult to implement in clinical practice . Severalpatients . The exaIndividual clinical trials and meta-analyses have shown that different fluid therapy regimens produce significantly different clinical outcomes and have resulted in considerable controversy as to the best approach. Table et al. compared the fluid therapy practices of both American Society of Anesthesiologists (ASA) and European Society of Anesthesiology (ESA) members [The use of standardized fluid therapy protocols within the perioperative space is limited despite strong evidence of benefit. A survey by Cannesson  members . StandarLack of standard criteria for fluid therapy results in significant clinical variability relative to the type and volume of fluid administered. This variability is linked to variable outcomes and makes it difficult to assess the effectiveness of different approaches ,63. A unBased on the abovementioned acknowledgements, a group of 72 researchers, well known within the field of fluid resuscitation, were invited, via email, to attend a meeting that was held in Chicago in 2011 to discuss perioperative fluid therapy. From the 72 invitees, 14 researchers representing 7 countries attended, and thus, the international Fluid Optimization Group (FOG) came into existence. These researches, working collaboratively, have reviewed the data from 162 different fluid resuscitation papers including both operative and intensive care unit populations.There was no human research involved with this manuscript.This manuscript is the result of 3 years of evidence-based, discussions, analysis, and synthesis of the currently known risks and benefits of individual fluids and the best methods for administering them. The results of this review paper provide an overview of the components of an effective perioperative fluid administration plan and address both the physiologic principles and outcomes of fluid administration.We present our evidence-based suggestions and individualized algorithms for a standardized approach to perioperative volume therapy for surgical patients. We propose specific recommendations for fluid administration which are organized into seven tenets as follows: 1) fluid responsiveness, dynamic indices, and the gray zone; 2) considerations of the composition of crystalloids and colloids; 3) evidence-based guidelines and individualized algorithms; 4) perioperative fluid plan; 5) goal-directed therapy; 6) the fluid challenge; and 7) maintenance fluids.Recommendations are italicized.R-R intervals and who are receiving controlled mechanical ventilation with tidal volumes between 8 and 10 ml/kg, fluid responsiveness is most effectively assessed using dynamic indices. These should be measured in a uniform manner before and promptly after each fluid intervention [In patients who have cardiac rhythms with regular rvention -69. Currrvention .The role of echocardiography, both transthoracic and transesophageal, can be critical when evaluating both fluid responsiveness and cardiac function. In addition, echocardiography is of particular use when assessing volume responsiveness in patients undergoing open chest surgery where the predictive ability of dynamic indices is also reduced .delta IVC and delta SVC diameter during positive pressure ventilation have shown to be effective for evaluating fluid responsiveness [Static parameters  derived from transesophagic echocardiography (TEE) monitoring are not useful in predicting volume responsiveness . On the siveness . As withWhile dynamic indices are excellent for predicting volume responsiveness, measured changes in cardiac output (∆CO) or stroke volume (∆SV) may be required to assure the effectiveness of a fluid bolus ,75. DynaDynamic indices have been repeatedly shown to accurately reflect fluid responsiveness and do so better than commonly used static hemodynamic parameters. These parameters have been validated and used to guide fluid therapy in a variety of surgical patients, including those undergoing major abdominal ,68,76-78per se is not always the correct therapy for hemodynamic instability.Dynamic parameters should be an integral part of GDT protocols for those patients in which they can be accurately measured. ∆CO or ∆SV can be used in the remaining patients. Not taking into account the status of fluid responsiveness when making fluid therapy decisions is bound to result in unjustified fluid administration even when GDT is being used. In addition, dynamic parameters may precede continuously measured CO, heart rate, and blood pressure in alerting to the development of hypovolemia and may therefore trigger an early and justified fluid administration ,92. It iet al. showed that, despite strong predictive value, there is a range of PPV values, named the gray zone (between 9% and 13%), for which fluid responsiveness cannot be reliably predicted in 25% of patients during general anesthesia [The predictive ability of various dynamic indices has been compared in a number of studies. PPV had been found to be somewhat more accurate than the SPV and SVV ,94,95. Hesthesia . Moreoveesthesia . Thus, westhesia .The interaction between PPV and SVV (PPV/SVV) has also been studied as a measure of dynamic vascular compliance ,98. ThesSince pulse oximetry is a standard noninvasive intraoperative monitor, the respiratory variation in the plethysmographic waveform (PWV) is potentially the most commonly available dynamic parameter in mechanically ventilated anesthetized patients . The majFluid responsiveness measures cannot be used in all patients and at all times in many patients. Dynamic indices have a high predictive value in determining fluid responsiveness; however, specific criteria must be met in order to use these indices to assess fluid responsiveness. Intraoperative motion, electrosurgical equipment, and physiologic artifact (noise) can interfere with the accurate interpretation of dynamic indices. Four primary limitations may exist in the use of dynamic indices. First, arrhythmias  preclude the use of SPV, PPV, SVV, and PWV to predict volume responsiveness, while inferior and superior vena cava variability remain accurate. The same limitation of SPV, PPV, SVV, and PWV is seen in subjects having varying levels of spontaneous inspiratory efforts. Again, inferior vena and superior vena cava diameter variability may remain predictive of volume responsiveness during spontaneous breathing. Second, if tidal volumes are <8 ml/kg, then the negative predictive value of SPV, PPV, SVV, and PWV is decreased whereas threshold values >13% variation still retain their positive predictive value. Third, marked decreases in chest wall compliance will decrease the positive predictive value of all indices whereas intra-abdominal hypertension may mask hypovolemia but will not alter the volume responsiveness prediction value of these indices. Fourth, in the setting of acute cor pulmonale, with marked ventricular interdependence, one will see a paradoxical positive SPV, PPV, SVV, or PWV which will increase more with fluid resuscitation. Thus, when dynamic indices are utilized for guiding fluid therapy, some measure of the effectiveness of augmented perfusion should be considered.Importantly, if these indices have values >20%, then the subject is clearly volume responsive. However, values from 9% to 13% may represent a ‘gray zone’ with less positive and negative predictive values and greater patient-to-patient variability. In these cases and when any of the above limitations precludes the use of these parameters, one may consider performing a fluid challenge or passive leg raising (PLR) maneuver . In contWe recommend that dynamic parameters be used as an integral part of GDT protocols. The limitations of each dynamic index must be taken into consideration as well as the concept of a gray zone. Dynamic parameters neither provide a measure of fluid bolus effectiveness nor should they to be used as an indication to give fluids. The final decision to administer fluids must be supported by the apparent need for hemodynamic improvement, the presence of fluid responsiveness, and the lack of associated risk.There has been extensive research evaluating the risks and benefits of specific types of fluids and developing alternative solutions that restore effective circulatory volume and enhance microcirculatory flow. Despite all these efforts, the ideal resuscitation fluid or combination of fluids remains undefined.Crystalloids are electrolyte solutions which are best used to replace extracellular volume losses from perspiration, respiration, and urine output. Although crystalloids increase vascular volume and may improve hemodynamics, the effectiveness is transient and less than colloid solutions. Crystalloids can be classified by their composition and osmolality. Normal saline (NS) is slightly hypertonic at 308 mOsm/l, and lactated Ringer’s (LR) is slightly hypotonic at 273 mOsm/l comparing to plasma osmolality. Plasmalyte is the most balanced isotonic electrolyte solution and has an osmolality of 294 mOsm  of colloid  for fluid resuscitation has been questioned ,107. Tabversus crystalloids on mortality in patients admitted to the ICU with hypovolemic shock. There was no difference in 28-day mortality between patients resuscitated with crystalloids or colloids. However, the 90-day mortality was significantly reduced in patients treated with colloids [versus crystalloid in 1,818 septic patients demonstrated that the colloid group had a higher mean arterial pressure during the first 7 days whereas there were no differences in the total amount of fluids administered between the two groups and both 28-day and 90-day mortality rates were similar. Thus, there is no compelling evidence that adding colloids to fluid resuscitation materially alters clinically relevant outcomes [The CRISTAL trial compared the effects of fluid resuscitation with colloids colloids . On the colloids ,113. Howoutcomes .versus standard of care therapy suggest benefit of nonstarch colloids such as albumin. It should be noted that the deleterious effects of starches have largely been reported in ICU trials where starch therapy was used for multiple days. In contrast, the beneficial effects of perioperative GDT trials that included starch-based volume therapy were only of limited duration and thus exposure. For this reason, we cannot conclude that the deleterious effects of starches shown in the ICU population are generalizable to the limited use that occurs in the immediate surgical space. Serious thought to the individual surgical patient’s co-morbidities, especially acute kidney injury, can inform the anesthesiologist of potential increased risk of starch-based colloid therapy.Given the evidence of harm and lack of significant clinical benefit in critically ill patients, when considering the administration of synthetic colloids, the anesthesiologist should first assess patient-specific risk. There is no evidence that the deleterious effects of starch-based colloids occur with albumin. The beneficial hemodynamic effects of colloid in GDT groups A recent Cochrane meta-analyses has concluded, however, that there is no evidence from randomized clinical trials that resuscitation with colloids, instead of crystalloids, reduces the risk of death in patients with trauma, burns, or following surgery . Common Similar caution and approach should be applied to other synthetic colloids such as dextran and gelatin. Scant clinical evidence exists as to either benefit or harm regarding to the administration of other colloid solutions such as dextran or gelatin to surgical patients. Siting theoretical safety concerns, some authors posit caution for the routine use of these fluids in surgical patients . It shouWe recommend crystalloid solutions for routine surgery of short duration. However, in major surgery, the use of a goal-directed fluid regimen containing colloid and balanced-salt solutions is recommended. Though a black box warning for the use of starch solutions exists within the US, there is limited data relative to their harm in the perioperative space. Careful consideration should occur in patients with known renal dysfunction and/or sepsis prior to administering starch solutions.Disagreement about optimal perioperative fluid therapy is exacerbated by the lack of uniform definitions for standard, restrictive, and supplemental fluid delivery . This, iGuidelines are general suggestions of care based on principles extracted from evidence-based findings and consensus. Algorithms are highly specific as to the variable(s) used, their target values, and their specific steps.versus the traditional care group [et al. demonstrated that significant reductions in complications and readmissions could be realized with the implementation of a simple two-step ERAS protocol focusing on early ambulation and alimentation following colorectal surgery [The difference between a guideline and an algorithm is important. Guidelines do not provide sufficient detail to reduce variation of care. Two anesthesiologists could strictly adhere to a guideline, but their specific fluid therapy delivered to an identical patient could be quite different. Even one of these individual anesthesiologist’s practices for two identical patients could differ from 1 day to another. Improved outcomes, reduced readmissions, and reduced costs have resulted when quality improvement programs have been implemented to reduce variability ,121. There group . ImportaImportantly, algorithms should not be fixed, they should allow for individualizing fluid therapy based on changing physiologic need and response to fluid and drug therapy. Algorithms can become very detailed and are likely to be best implemented with computerized decision support ,126. BotComputerized decision support and implementation of closed-loop fluid administration has been described. Significant regulatory challenges exist before these systems can be introduced into clinical practice ,128. RecWe recommend the use of algorithms as part of the perioperative fluid plan. These should be available and easily accessible within all operating rooms. We encourage continued development, refinement, and testing of computerized decision support tools.Patient status ;Surgical risk ;Selection of hemodynamic monitoring based upon patient and surgical risk as well as the anesthesiologists’ clinical management needs . Figure The use of protocols for perioperative hemodynamic support which enhance tissue perfusion has been shown in multiple meta-analyses to reduce organ dysfunction, mortality, and HLOS ,130,131.et al. [Vincent et al.  proposedet al. ,134. AdvIn low-risk patients and low-risk surgery, the use of ASA standard monitors is often sufficient. However, if the associated risk or surgical procedure escalates, or if unexpected patient instability develops, additional expertise and/or monitoring are requisite. The advancement of monitoring is not without increased risk and cost; thus, these tools should only be applied when needed to provide a means to better detect and treat tissue mal-perfusion and potential organ dysfunction. Fluid therapy is a cornerstone for perioperative medicine, but clarity on when not to infuse fluids is as important as when to infuse. Hemodynamic and other advanced monitoring is often the best means to assess and assure the optimization of intravascular volume, pressure, perfusion, and oxygenation.Invasive cardiovascular monitors can be considered in patients in whom tight hemodynamic control is needed to prevent rapid organ deterioration, for example, significant heart or brain disease and in high-risk surgical cases, for example, aortic and heart surgery. Indices from commonly used invasive monitors include intra-arterial pressure from an arterial catheter, right and left heart filling pressures, and central or mixed venous oxygenation saturations from central venous or pulmonary artery catheters, respectively.There are several commercially available noninvasive or minimally invasive technologies that employ arterial pulse contour analysis, bioimpedance, or bioreactance and which provide continuous cardiac output, dynamic indices, and systemic vascular resistance. Finally, echocardiography, transthoracic and transesophageal (TEE), is becoming increasingly utilized in at risk patients. Mastering cardiac performance and volume assessment by TEE may improve currently available GDT algorithms.We recommend that a perioperative fluid plan be developed by each department, facility, or health system and used by all anesthesiologists. Clinical needs, invasiveness, accuracy, and precision of available technologies should be considered when selecting monitoring devices.Until recently, perioperative fluid replacement was guided primarily by estimates of known or anticipated fluid deficits and replacing these using fixed calculations for administration of intravenous fluid. Little outcome data exist that support the widespread use of fixed perioperative fluid regimens. Recent approaches have focused attention on the type of surgery being performed and the impact of the following outcomes: 1) the type of fluid being administered; 2) the timing of fluid administration; 3) the rate of fluid administration ,136; 4) Effective fluid therapy algorithms incorporate GDT. Table There are two overall challenges for fluid optimization as follows: 1) how to best identify hypovolemia and tissue hypoperfusion; and 2) how to best optimize vascular volume, cardiac filling, global, and regional perfusion and tissue oxygenation.fluid responsiveness, that is, a measure of the change in CO in response to an increase in preload [The most common parameters that are used to guide the need for hemodynamic support and perioperative fluids include clinical experience, urinary output, mean arterial pressure, and CVP ,137. Oth preload  as discu preload .Although the goal-directed fluid therapy concept was first suggested more than 30 years ago , there rWhile the benefits of perioperative goal-directed fluid therapy have yet to be proven, the bulk of clinical research supports the implementation of a two-step GDT plan which is to begin immediately after induction of anesthesia. First, determine if the patient requires hemodynamic support or augmentation of cardiovascular function. Second, if the need is apparent and the patient is fluid responsive, fluid bolus therapy should be considered and guided by continual, and if available continuous, assessment of fluid responsiveness as described below.A fluid challenge is one of the best tools that the anesthesiologist has for assessing fluid responsiveness. To test fluid responsiveness, a change in preload (fluid bolus) must be induced while monitoring the subsequent change in stroke volume, cardiac output, and dynamic indices .a means to assess the patient’s response to fluid with changes in dynamic indices and static indices of volume, flow, and oxygenation;a prompt increase in intravascular volume and usually a needed improvement in flow (cardiac output).The use of a fluid bolus provides two advantages as follows:A fluid bolus is a provocative test of the circulation, similar to the use of a step function in engineering to define a system. The use of a ‘test’ that uses a small amount of fluid (bolus) to assess the volume responsiveness may reduce the risk of a too liberal fluid strategy and the possible consequences of fluid overload. These tools help to determine the requirements for additional fluid therapy avoiding the deleterious consequences of fluid overload through its small volume and targeted administration .It is important to stress that the fluid challenge technique is a test of the cardiovascular system. It allows clinicians to assess whether a patient has enough preload reserve to increase stroke volume with further fluids. Fluid therapy should be considered after a positive response to a fluid challenge. In contrast to a single fluid challenge, fluids can also be infused in a controlled fashion based on an algorithm by repeating the fluid challenge as long as there is a positive response. This controlled approach is called stroke volume maximization and is the cornerstone of most goal-directed therapy protocols . Thus, tA fluid challenge should comprise four separate orders: the type of fluid to be infused, the volume of fluid to be infused, the rate of the infusion, and the stopping rules if untoward effects are seen before the full amount of the bolus is infused. For rapid infusions of very small boluses of fluid , stopping rules are probably not necessary. But if larger amounts of fluids or longer infusion times are used, clear stopping rules are important to prevent right heart failure or pulmonary edema.Although no consensus is available for the type and exact dosing of fluid administration, boluses are best delivered at a rapid rate (5 to 10 min) with prompt assessment of the physiologic response. The magnitude of this response helps to determine the effectiveness of the fluid challenge as well as the requirements for additional fluid therapy. Taken together, this approach avoids the deleterious consequences of fluid overload . The peaEstablishing volume status is complex, making accurate prediction of an increase in stroke volume upon fluid load challenging. However, under conditions of hypovolemia and inadequate perfusion, there is greater vascular retention of infused volume due to physiologic compensatory mechanisms that act to maintain normal volume, pressure, and perfusion. These compensatory mechanisms include the renal response to elevated vasopressin, angiotensin, and aldosterone; reduced capillary filtration due to reduced venous and capillary pressures; and decreased capillary hydraulic conductivity due to fluid composition and decreased levels of atrial natriuretic peptide (ANP) ,150. TheWe recommend bolus therapy rather than continuous infusion when the goal is to improve pressure, perfusion, and oxygen delivery. Standardization of the fluid bolus relative to fluid composition, volume, infusion rate, and time to post bolus assessment should be implemented. The variables used for assessing the effectiveness of the fluid bolus should include appropriate changes in cardiac output or stroke volume.Traditional perioperative fluid administration is guided by estimates of both the preoperative fluid deficit and by ongoing sensible and insensible intraoperative fluid losses. The notion that all surgical patients are hypovolemic due to prolonged fasting, bowel preparation, and ongoing losses from perspiration and urinary output is unfounded. Preoperative volume status is typically unknown and should not be presumed to be either adequate or inadequate. Blood volume varies considerably between patients depending on gender, weight, and oxygen consumption -153. MorThe majority of the patients present with a minor functional intravascular deficit before surgery (200 to 600 ml) that is unlikely to have clinical significance . This maIn the clinical context of ambulatory surgery in low-risk patients, a more liberal fluid strategy may be beneficial. Up to 20 to 30 ml/kg/h of crystalloid infusion reduces postoperative dizziness, drowsiness, pain, nausea, vomiting, and hospital length of stay -160. To We recommend that maintenance fluids be administered at a rate of 1 to 2 ml/kg/h for patients undergoing procedures of longer duration or magnitude. Patients undergoing outpatient procedures may benefit from higher maintenance fluid rates.Although perioperative fluid management remains a highly debated subject, data suggests that goal-directed fluid therapy with the objective of hemodynamic optimization can reduce complications after major surgery. Specific hemodynamic goals include maintaining adequate circulating volume, perfusion pressure, and oxygen delivery. Lack of standard criteria for perioperative fluid therapy results in significant clinical variability relative to its administration.In summary, fluids should be treated as any other intravenous drug therapy, and thus, careful consideration of its timing and dose is mandatory. A perioperative fluid plan should be developed which is easily understood and used by all anesthesiologists within a group, facility, or healthcare system. Determining both the need for augmented perfusion and fluid responsiveness is fundamental when making fluid therapy decisions to avoid unjustified fluid administration. Balanced crystalloid solutions should be given for short duration/low-risk surgical patients. Procedures of higher complexity are best managed with a combination of crystalloid and colloid therapy. When considering the administration of starch containing solutions, the anesthesiologist should first assess patient-specific risk. Finally, we recommend the use of algorithms as part of the perioperative fluid plan.


Difficulties in the highly sensitive detection of tumour microfoci represent a critical obstacle toward improved surgical intervention in liver cancer. Conventional preoperative imaging methods and surgeons’ subjective experience are limited by their inability to effectively detect tumour lesions measuring less than 2 mm; however, intraoperative fluorescence molecular imaging may overcome this limitation. Here, we synthesised an arginine-glycine-aspartic acid (RGD)-conjugated mesoporous silica nanoparticle (MSN) highly loaded with indocyanine green (ICG) dye that could accurately delineate liver cancer margins and provide excellent tumour-to-normal tissue contrast intraoperatively. The increased ICG loading capacity and tumour specificity enabled the identification of residual microtumours and satellite lesions measuring less than 1 mm in living mice. Histological analysis validated the sensitivity and accuracy of this approach. We believe this technique utilising a new fluorescent nanoprobe with intraoperative optical imaging may offer a more sensitive and accurate method for liver cancer resection guidance, resulting in better surgical outcomes. The outcomes of liver cancer surgeries rely heavily on the accurate localisation of tumour lesions12457891112414One strategy for obtaining better TBR in hepatic tumour sites has been to increase the tumour specificity of the fluorescent probe16171882021Mesoporous silica nanoparticles (MSNs) have received significant attention as drug nanocarriers in cancer therapy due to their unique properties, including high chemical stability, controllable pore size, large surface area, and superb drug loading ability232426vβ3 receptor that is overexpressed in many tumour types28In this study, we synthesised arginine-glycine-aspartic acid (RGD)-conjugated highly loaded indocyanine green (ICG) MSNs (ICG/MSNs-RGD) as a fluorescent probe and explored its application to intraoperative NIR-image-guided liver tumour resection. RGD is a peptide ligand for the integrin αin vivo, and particles with sizes between 30 and 150 nm are particularly favorableThe size and morphology of a nanoprobe are critical factors affecting tumour targeting In vitro cytotoxicity testing of the nanoparticles indicated that it has good biocompatibility -fLuc tumours with high integrin αvβ3 expression. Two groups of three mice were injected intravenously (i.v.) with either the ICG/MSNs or ICG/MSNs-RGD probe . Continuous fluorescence imaging over 120 h indicated marked accumulation of both probes in tumour lesions up to 96 h; however, the ICG/MSNs-RGD probe displayed better optical contrast than the ICG/MSNs in the tumour region from the 24 h time point onward  xenografts. The tumour margins were very difficult to visually identify after removing the skin ; howeverin vivo , as wellin vivo . To furtin vivo . The tumin vivo .Quantitatively, NIR imaging showed a notable improvement in the TBR over that of GFP  to completely remove the tumour lesion while minimising surgical trauma. NIR and GFP fluorescence images were obtained before and after the resection to verify tumour residuals . HoweverHaving established the efficacy of intraoperative NIR-FMI with ICG/MSNs-RGD, we sought to investigate the performance of this approach for the detection of liver cancer metastases and satellite foci. Intraoperative NIR-FMI-guided surgery was performed on three mice with intrahepatic metastases 48 h after the i.v. administration of ICG/MSNs-RGD . Notably, only the implanted tumours (5.09 ± 2.31 mm) could be visually recognised based on the surgeons’ subjective experience after exposing the liver , and NIRUnexpectedly, we found five other mice with peritoneal satellite lesions, likely due to the leakage of tumour cells during implantation. The average size of these lesions in the peritoneum was 1.32 ± 0.27 mm , whereasAs the first capillary bed to be encountered by circulating malignant cells, the liver commonly contains metastases from primary tumours in other organs. Therefore, we also simulated liver metastasis from breast cancer, which is the second most common hepatic metastasis in clinical practice3132The effectiveness of intraoperative fluorescence imaging guidance has been validated in various preclinical and clinical applications3435373839vβ3-overexpressed tumour cells, and its specificity was verified both in vitro and in vivo. With the specific binding of the RGD peptide to integrin αvβ3, the nanomaterials could be internalised into cells by clathrin-mediated endocytosis, also known as receptor-mediated endocytosis (RME). Several studies have reported that RGD-containing nanomaterials could enhance cellular uptake through the RME pathway411729in vivo administration demonstrated its biodistribution, clearance, and elimination.The RGD-modified probe targeted integrin αIn addition to these demonstrated advantages, ICG/MSNs-RGD was specifically designed to offer sufficient optical contrast for tumour microfoci, despite the limitations of probe uptake by small tumour lesions. This was achieved by employing MSNs as nanocarriers that encapsulated the ICG; for each successfully delivered ICG/MSNs-RGD nanoparticle, multiple ICG molecules could be effectively activated and contribute to the optical contrast between tumour and normal tissues. After adjusting the MSN mesoporosity to 5 nm, the encapsulation efficiency reached an extraordinary 37.5% . The hig2424To prove our hypothesis, we established different liver cancer-bearing mouse models and applied intraoperative NIR-FMI with ICG/MSNs-RGD to simulate various clinical scenarios. The experiments conducted by the two surgeons demonstrated that the probe facilitated effective detection of liver cancer residuals less than 1 mm in size. The tumour metastasis experiment also indicated that a surgeon can easily recognise intrahepatic and extrahepatic liver cancer satellites of less than 1 mm using the newly developed probe. This is likely the most sensitive intraoperative NIR-FMI performance in liver cancer surgery to date. Moreover, ICG/MSNs-RGD provided TBR values for the liver microtumours that exceeded 4.5, which revealed the superb optical contrast between the tumours and surrounding tissues. These experiments proved that ICG/MSNs-RGD could provide extremely high sensitivity for the intraoperative detection of tumour foci at the micrometre level, while maintaining a field of view encompassing more than half of the body of the mouse.in vivo NIR imaging studies, the optical signal penetration depth is the major limitation of this technique, which constrains effective liver tumour detection to superficial lesions21vβ3. However, dual-targeted nanoparticles may be a powerful tool for overcoming these problems. A recent report described a dual-targeted probe (cyclic-RGD-PLGC(Me)AG-ACPP) that could be taken up by multiple types of human cancer cells and delivered more therapeutic or diagnostic agents to tumour regionsAs in other ICG/MSNs-RGD can greatly assist surgeons in the intraoperative detection of liver cancer. Accurate delineation of tumour borders and sensitive detection of tumour residuals are both benefits of the high ICG loading capacity of the MSNs. This molecular design strategy can be extended to other tumour imaging applications. Through the conjugation of different targeting domains (peptides or antibodies), this method has great potential in the highly sensitive and accurate visualisation of tumour microfoci with expression of various biomarkers3vβ3 receptor targeting specificity. The probe demonstrated precise tumour margin delineation and highly sensitive microfoci detection during liver cancer surgery. Both tumour metastases and surgical residuals at the micrometre level were effectively captured intraoperatively with a macroscopic field of view. Furthermore, the probe exhibited promising biocompatibility, good optical stability, and high water dispersibility. We believe this novel fluorescent nanoprobe with the new high-fluorochrome-loading design strategy will afford more sensitive and accurate guidance of liver cancer resection to achieve better surgical outcomes.In this study, we successfully constructed an active tumour-selective probe, ICG/MSNs-RGD, using MSNs with a high ICG-loading ability and high α2-SiO2 was collected, extensively washed with ultrapure water and absolute ethyl alcohol, and then dried under high vacuum at 30 °C overnight to remove any remaining solvent.MSNs with customisable pore and particle sizes were generated by hydrothermal synthesis242-SiO2 (10 mg) to ICG solution  and vibrating overnight  and SEM  were used to evaluate the morphology and size of the MSNs and ICG/MSNs-RGD, with a 100 kV acceleration voltage. The hydrodynamic particle size distribution was measured by dynamic light scattering (DLS) using a Malvern Zetasizer . The fluorescence excitation and emission spectra of the ICG and ICG/MSNs-RGD were measured with a fluorescence spectrofluorometer . For stability analysis, the ICG/MSNs-RGD (50 μg) was added to FBS  and its optical absorbance at 780 nm was monitored by UV-Vis spectroscopy  at multiple time points for 24 h. A serum-only control solution was also monitored.2. Cells were seeded into 96-well microtiter plates at 10,000 cells per well. After incubation for 24 h, the cells were washed with phosphate-buffered saline (PBS), and the medium was replaced with fresh medium containing various concentrations of the MSNs or ICG/MSNs-RGD . An equivalent volume of fresh medium (200 μL/well) was added to the control wells. After 24 h incubation, the treated cells were washed with PBS, and then fresh medium (100 μL) and MTT solution  were added to each well and cultured for another 4 h. The old medium was removed, and dimethyl sulfoxide  was added to each well. The absorbance at 490 nm was read using a microplate reader to calculate relative cell viabilities.The cellular cytotoxicity of the MSNs and ICG/MSNs-RGD was assessed by 3--2,5-diphenyltetrazolium bromide (MTT) assay . GFP-transfected luciferase-expressing human hepatocellular carcinoma cells (Hep-G2-GFP-fLuc cells), which expressed αvβ3 receptors and MDA-MB-231-fLuc human mammary cancer cells were purchased from the Academy of Military Medical Sciences (China) and cultured in Dulbecco’s modified Eagle’s medium  supplemented with 10% FBS and 1% penicillin/streptomycin (Promega) at 37 °C in an atmosphere containing 5% COvβ3 receptor, we used MDA-MB-231-fLuc and MCF-7  cells, which exhibited high and low integrin αvβ3 expression, respectively. For this, 105 cells were cultured in 96-well microtiter plates for 24 h prior to the addition of ICG, ICG/MSNs, or ICG/MSNs-RGD (the ICG concentration was 5 μg/mL) for 3 h. After washing three times with cell culture medium and twice with PBS, culture medium (100 μL) was added to each well, and the absorbance at 780 nm was measured with a microplate reader.To evaluate specificity for the integrin αMDA-MB-231-fLuc and MCF-7 cells were incubated with ICG/MSNs-RGD (0.2 mg/mL) for 3 h. The cells were washed three times with cell culture medium and twice with PBS, and then additional cell culture medium (2 mL) was added for continuation of cell culture. A split fluorescence microscope and NIR-fluorescence imaging-guided surgery system were used to observe the cells at excitation and emission wavelengths of 780 and 840 nm, respectively.All animals were purchased from the Department of Experimental Animals at Peking University Health Science Center (China). All experimental protocols were approved by the Institutional Animal Care and Ethics Committee of Zhujiang Hospital of Southern Medical University, and all the methods were carried out in accordance with the approved guidelines.6 Hep-G2-GFP-fLuc cells into the left upper elbow/back region in 6–7-week-old BALB/c nu/nu male mice.A subcutaneous hepatic tumour model was established by subcutaneous injection of 2 × 106 Hep-G2-GFP-fLuc cells into the livers of 6–7-week-old male BALB/c nu/nu mice in a vehicle of Matrigel  in PBS with a 50-μL injection volume. Tumours were allowed to grow for 12 days, and then visualised with an IVIS Spectrum Imaging System . For imaging, mice were anesthetised with a 3% isoflurane /air mixture, injected with luciferin solution , and then imaged 8 min later in the supine position.An orthotopic hepatic cancer model was established by injecting 2 × 106 MDA-MB-231-fLuc cells were injected into 6–7-week-old female BALB/c nu/nu mice in a vehicle of Matrigel (4 mg/mL) in PBS with a 50-μL injection volume. Tumours were allowed to grow for 12 days prior to monitoring with IVIS as described above.For the metastatic breast cancer model, 2 × 10in vivo fluorescence imaging, anesthetised with 3% isoflurane/air, and treated with ICG/MSNs or ICG/MSNs-RGD  via tail vein injection. Mice were monitored by IVIS at 10 min, 24 h, 48 h, 72 h, 96 h, and 120 h postinjection at excitation and emission wavelengths of 780 and 840 nm, respectively. Imaging data were analysed with IVIS Living Image 3.0 software (PerkinElmer).Six mice with subcutaneous hepatic tumours measuring more than 5 mm in diameter were selected for Surgical studies were performed by two general surgeons . Twelve mice (six mice per surgeon) were treated with ICG/MSNs-RGD  48 h prior to surgery. Surgeons were asked to remove tumour tissue with maximum protection of the normal tissue based on visual inspection and palpation. After this procedure, NIR and GFP images were acquired to determine whether residual tumour tissues were present. If tumour tissue remained, the surgeons continued the resection with imaging guidance until all tumour tissue was removed. Tumour numbers and sizes were recorded, and the resected tissue was histologically examined.2) within the tumour nodules and adjacent liver tissue and measuring their average fluorescence intensity. The TBR value was then calculated as the average fluorescence intensity of the tumour nodule divided by that of surrounding areas.Through the continuous monitoring of 20 mice with orthotopically implanted liver tumours (Hep-G2-GFP-fLuc cells) using bioluminescence imaging, we found intrahepatic metastases in three mice and peritoneal satellite lesions in five other mice. Surgical studies were performed by a general surgeon on animals administered ICG/MSNs-RGD  48 h prior to surgery. Both orthotopic and metastatic liver cancer model mice were anesthetised with 4% chloral hydrate  solution and fixed on the operational bed in the supine position. The abdominal cavities were exposed with a longitudinal incision in the middle of the abdomen. The surgeon then resected all the primary and metastatic tumour tissue under the guidance of the NIR-fluorescence imaging system. The TBR ratio for each ICG/MSNs-RGD probe was evaluated by drawing 10 regions of interest  compound , snap-frozen in liquid nitrogen, and stored at –80 °C. Sections were cut on a cryostat microtome (Leica) at a thickness of 8 μm and stained with H&E. Following pathological examination, ICG/MSNs-RGD-based fluorescence images were collected using an NIR-fluorescence imaging system to determine the fluorescence signal differences between tumour tissues and normal tissues.p values of less than 0.05 were considered significant.Statistical analysis was performed using SPSS v.20 . Data are expressed as the means ± standard deviations (SDs). Two-tailed, independent two-sample t-tests were used to assess differences in the optical densities and fluorescence intensities between groups. Differences with How to cite this article: Zeng, C. et al. Intraoperative Identification of Liver Cancer Microfoci Using a Targeted Near-Infrared Fluorescent Probe for Imaging-Guided Surgery. Sci. Rep. 6, 21959; doi: 10.1038/srep21959 (2016).


Caenorhabditis elegans (C. elegans) as a model to study the mechanism underlying insulin regulation of GLUT, we identified that FGT‐1 is most likely the only functional GLUT homolog in C. elegans and is ubiquitously expressed. The FGT‐1‐mediated glucose uptake was almost completely defective in insulin/IGF‐like signaling (IIS) mutants daf‐2 and age‐1, and this defect mainly resulted from the down‐regulated FGT‐1 protein expression. However, glycosylation may also be involved because OGA‐1, an O‐GlcNAcase, was essential for the function of FGT‐1. Thus, our study showed that C. elegans can be a new powerful model system to study insulin regulation of GLUT.Insulin signaling plays a central role in the regulation of facilitative glucose transporters (GLUTs) in humans. To establish  When blood glucose levels increase after meals, insulin is secreted to activate glucose uptake and utilization in peripheral tissues, mainly muscle and adipose tissues Caenorhabditis elegans (C. elegans). In C. elegans, most of the major signaling molecules in the insulin cascade are well conserved, although insulin‐like growth factors (IGF) may function through the same cascade [termed insulin/IGF‐like signaling (IIS)] unlike in humans, and IIS has been known to play important functions in many physiological processes, such as reproduction, growth, metabolism, and aging C. elegansC. elegans may be an attractive model system to study the mechanisms underlying insulin regulation of glucose transport.Insulin signaling is highly conserved in the animal kingdom, including C. elegans as a model system for the study of insulin regulation of GLUT functions, we characterized nine GLUT candidates in C. elegans (FGT) based on their high sequence homologies to human GLUTs in our previous study Xenopus oocytes. The localization of FGT‐1 was mainly restricted to the digestive tract when expressed under its 2 kb promoter sequence C. elegans.To explore In this study, we investigated the tissue localization of another FGT‐1 alternative splicing isoform, FGT‐1B, and re‐examined the tissue localization of FGT‐1A in response to a longer promoter sequence. We also showed that FGT‐1 is regulated by IIS by comparing the differences in glucose uptake between FGT‐1 mutants in wild‐type (wt) or IIS mutant backgrounds. We further investigated the regulatory mechanisms of IIS on FGT‐1 function.C. elegans strains used in this study are described in Data S1. The C. elegans strains were cultivated at 20 °C under standard conditions unless otherwise specified All plasmids and fgt‐1a were generated in our previous study fgt‐1b, fgt‐1a(tm3165), and fgt‐1b(tm3165) were synthesized by in vitro transcription from pSP‐fgt1b, pSP‐fgt1atm, and pSP‐fgt1btm, respectively, using the mMessage mMachine kit . The 2DG uptake analysis of wt or mutated FGT‐1A and FGT‐1B was performed in Xenopus oocytes as described previously The cRNAs of m 2DG containing 3 μCi 3H‐2DG in M9 saline in the presence or absence of 100 μm phloretin or phlorizin. The worms were incubated in the uptake solutions for 2 h at 20 °C and then washed thoroughly three times with ice‐cold PBS containing 0.5% Tween‐20 prior to lysis in 0.5% SDS containing 60 μg·mL−1 proteinase K for 1 h at 55 °C. The radioactivity of the lysed worms was counted using a Tri‐Carb Liquid Scintillation Counter 2900TR . Each experiment was performed independently four times.Synchronized young adult worms were washed out from the culture plates and incubated in M9 saline for 1 h. A 10% volume of worms was collected for protein quantitation. The remaining worms were subjected to the uptake assay using 0.5 mRNAi feeder plasmids of DAF‐2 and DAF‐16 were obtained from Addgene . An RNAi feeder plasmid of AGE‐1 was obtained from Source Bioscience . RNAi feeder plasmids of AKT‐1 and OGA‐1 were obtained from GE Healthcare Dharmacon Inc . These plasmids were transformed into HT115 (DE3) bacteria, and RNAi was performed by culturing the worms on plates together with these feeding bacteria.image lab 4.1  and normalized to the level of β‐actin.The mRNA levels of FGT‐1, DAF‐2, AGE‐1, AKT‐1, DAF‐16, and OGA‐1 were measured by reverse transcription quantitative PCR (RT‐qPCR) with primer pairs FGT1q, DAF2q, AGE1q, AKT1q, DAF16q, and OGA1q, respectively (Table S1) fgt‐1a or fgt‐1b cRNA‐injected oocytes less than three times the mean value of water‐injected oocytes was considered as injection failure and discarded from the analysis. The 2DG uptake in fgt‐1a or fgt‐1b cRNA‐injected oocytes was corrected by subtraction of the mean 2DG uptake of water‐injected oocytes.For the 2DG transport assay in oocytes, any uptake in graphpad prism 6.03  and jmp pro 11.2 .The statistical analysis of the individual experiments is indicated in the figure legends, and the analyses were conducted using C. elegans with glucose transport activity and its expression was mainly observed in the digestive tract in our previous study , we hypothesized that another FGT‐1‐splicing isoform, FGT‐1B, is expressed in other tissues. FGT‐1B utilizes a distinct exon 1 from FGT‐1A and, therefore, has a slightly different promoter sequence  did not differ from that of FGT‐1A as reported previously Because FGT‐1(A) was identified as the sole GLUT homolog in C. elegans, and that the 2 kb promoter sequence may not be long enough to include all of the regulatory elements of the fgt‐1 promoter.To examine whether the 2 kb upstream sequence of FGT‐1 faithfully represents the endogenous localization of FGT‐1, we reanalyzed the tissue localization of FGT‐1A::GFP driven by a longer 5 kb promoter sequence. As shown in Fig. C. elegans, we examined the activities of facilitated glucose transport and Na+‐dependent glucose transport in wt and IIS mutant animals by measuring the uptake activities of tritium‐labeled 2DG (3H‐2DG) in wt and mutant strains of fgt‐1(tm3165), daf‐2(e1370), fgt‐1(tm3165); daf‐2(e1370), age‐1(hx546), akt‐1(mg306), and daf‐16(mgDf50); daf‐2(e1370) in the presence or absence of phloretin, a facilitated glucose transport inhibitor, or phlorizin, a Na+‐dependent glucose transport inhibitor  mutant was confirmed in Fig. fgt‐1, daf‐2, fgt‐1; daf‐2, and age‐1 mutants were similar and approximately 40% lower than those in wt (P < 0.001). The akt‐1 mutant also displayed lower glucose uptake than wt (P < 0.05) but higher uptake than the fgt‐1, daf‐2, fgt‐1; daf‐2, and age‐1 mutants (P < 0.05). In addition, glucose uptake activity in daf‐16; daf‐2 did not differ from that in wt  but not in the fgt‐1 and fgt‐1; daf‐2 mutants. Phloretin tended to inhibit 2DG uptake in age‐1 (P = 0.058)   and akt‐1(mg306) compared to wt, it was approximately 50% higher in the daf‐2(e1370) and daf‐16(mgDf50); daf‐2(e1370) mutants (P < 0.001)  but not by akt‐1 and daf‐16; daf‐2 RNAi  O‐linked glycosylation sites (NetOGlyc 4.0 Server). In C. elegans, OGT‐1 and OGA‐1 encode the sole O‐GlcNAc transferase and O‐GlcNAcase, respectively C. elegans, we first assessed 2DG uptake activity in the ogt‐1(ok1474) and oga‐1(ok1207) mutant strains. Although the ogt‐1 mutant showed the same level of 2DG uptake activity as wt, the transport activity was sharply decreased in the oga‐1 mutant (P < 0.001)  but not in the oga‐1 mutant ; oga‐1(ok1207) showed the same level of uptake activity as the oga‐1(ok1207) mutant, which suggested that the impaired 2DG uptake in the oga‐1 mutant resulted from the impaired function of FGT‐1. To examine this possibility, the mRNA and protein levels of FGT‐1 were analyzed in oga‐1 mutant or knockdown animals. There were no differences in FGT‐1 mRNA levels between wt and oga‐1 mutant strains, as measured by real‐time RT‐qPCR  in FGT‐1::GFP transgenic worms  In contrast to wt, the impaired 2DG uptake in the FGT‐1 null function mutant fgt‐1(tm3165) could not be further inhibited by phloretin, a GLUT inhibitor +/glucose cotransporter (SGLT) inhibitor fgt‐1 promoter, FGT‐1 was ubiquitously expressed in C. elegans  did not reveal any proteins with significant sequence homology to SGLTs. Thus, the transporters responsible for Na+‐dependent glucose transport in C. elegans remain to be identified.In our previous study et al. daf‐2 mutant animals, 2DG uptake activity was decreased to a level similar to that in fgt‐1 mutant animals because the activity did not further decrease in the fgt‐1; daf‐2 double mutant and could be inhibited only slightly by phloretin. These results indicated that FGT‐1‐mediated glucose uptake was almost completely impaired in the daf‐2 mutant. In addition, our data showed that the impaired function of FGT‐1 in the daf‐2 mutant was dependent on the DAF‐16 transcription factor because the DAF‐16 mutation in the daf‐2 mutant completely recovered the decreased 2DG uptake  We establish C. elegans as a potential unique model to study the mechanisms of insulin regulation of glucose transport because of its relative ease of genetic manipulation and (b) C. elegans may be used as a drug‐screening model to search for compounds that stimulate glucose uptake in patients with diabetes.In this study, we demonstrated that in SK designed and performed experiments and wrote the manuscript; ADM provided equipment and revised the manuscript; F‐QZ designed experiments and revised the manuscript.Data S1. Materials and methods.Table S1. List of primer sets.Click here for additional data file.


In patients with aneurysmal subarachnoid hemorrhage (SAH), hypervolemic therapy may result in fluid overload that may be associated with adverse clinical outcomes ,2. We hyFollowing the introduction of the hemodynamic protocol in 2011, 26 consecutive patients with SAH were included until 2013 Using TPT , cardiac output (CO), global end- diastolic volume index (GEDVI) and extravascular lung water index (EVLWI) were determined. Fluid administration was targeted at fluid unresponsiveness. Indications for start of the protocol were: hypotension (in spite of fluids), pulmonary edema or cardiac stunning, daily fluid balance ≤-1 l, cerebral ischemia (DCI) with progressive symptoms. Data were collected on fluid intake and output up to 3 days before and 3 days after the start of TPT. We assessed the course of fluid input and output and hemodynamic parameters before and after the start of the protocol with the generalized estimating equation.P < 0.001; day 0: +0.68 ± 0.29 l, P = 0.019; day 1: +0.73 ± 0.31 l, P = 0.02), increased fluid output , and consequently a strong decrease in fluid balance . The decreased fluid intake and fluid balances did not result in decreased CO, GEDVI or EVLWI on days 1 to 3 compared with day 0 .Mean age was 55 ± 16, and median Glasgow Coma Scale on admission was 8 (IQR 6 to 13). TPT was started at a median of 1 day after ICU admission (IQR 0 to 4). DCI developed in 70% and the in-hospital death rate was 45%. Compared with days preceding the protocol (day -3 to -1), TPT (day 0 to 3) was associated with decreased fluid intake (compared with day 3 as reference; day -1: +1.14 ± 0.31 l, Our data suggest that in poor-grade SAH patients goal- directed fluid management with TPT is feasible to decrease intake and increase diuresis without adverse effects on cardiac output or preload parameters. Future research should assess the effect of such a protocol on clinical outcomes.