Immunoglobulin G (IgG) effector functions are regulated by the composition of glycans attached to a conserved N-glycosylation site in the Fc part. Intrathecal production of IgG, especially IgG1, is a hallmark of multiple sclerosis (MS), but nothing is known about IgG Fc glycosylation in MS and in cerebrospinal fluid (CSF) in general.p values were adjusted for multiple testing.We applied mass spectrometry of tryptic Fc glycopeptides to analyze IgG Fc glycosylation ) in 48 paired CSF and serum samples from adult patients with MS or a first demyelinating event highly suggestive of MS (designated as MS cases), and from healthy volunteers and patients with other non-inflammatory diseases (control group). Our experiments revealed four main results. First, IgG1 glycosylation patterns were different in CSF vs. serum, in the MS group and even in control donors without intrathecal IgG synthesis. Second, in MS patients vs. controls, IgG1 glycosylation patterns were altered in CSF, but not in serum. Specifically, in CSF from the MS group, bisecting GlcNAc were elevated, and afucosylation and galactosylation were reduced. Elevated bisecting GlcNAc and reduced galactosylation are known to enhance IgG effector functions. Third, hypothesis-free regression analysis revealed that alterations of afucosylation and bisecting GlcNAc in CSF from MS cases peaked 2–3 months after the last relapse. Fourth, CSF IgG1 glycosylation correlated with the degree of intrathecal IgG synthesis and CSF cell count.The CNS compartment as well as the inflammatory milieu in MS affect IgG1 Fc glycosylation. In MS, the CSF IgG1 glycosylation has features that enhance Fc effector functions.The online version of this article (doi:10.1186/s12974-015-0450-1) contains supplementary material, which is available to authorized users. Thervy chain , 9.The functional in vivo relevance of different IgG Fc glycosylation patterns has been shown in animal models of systemic autoimmune diseases –14. The Glycosylation of IgG in blood is altered in human autoimmune diseases: in serum from patients with systemic autoimmune diseases such as rheumatoid arthritis, Lambert-Eaton myasthenic syndrome, and Guillain-Barré-Syndrome –27, IgG 35–55 induced experimental autoimmune encephalomyelitis as a model of MS [The translation into a therapeutic in vivo approach has been pioneered using the IgG-specific endoglycosidase EndoS : in vivoel of MS , as wellel of MS , 35, andel of MS . The regel of MS , 37.We present the first analysis of IgG glycosylation in the cerebrospinal fluid (CSF) and in MS. We applied mass spectrometry to determine IgG Fc glycosylation in paired CSF/serum samples and addressed the following questions: (1) Does IgG1 glycosylation differ between CSF and serum in the absence of inflammation? (2) Is multiple sclerosis associated with an altered pattern of IgG1 glycosylation in CSF or serum? (3) Is the pattern of CSF IgG1 glycosylation associated with time since last relapse and markers of inflammation in the CSF such as cell count and intrathecal IgG synthesis?n = 10), relapsing-remitting MS , secondary progressive MS , primary progressive MS ; all summarized as MS group). Even though the 2010 McDonald criteria allow for an earlier diagnosis, we adhered to the 2005 criteria since original MRI images were not available for all MS cases for reassessment. Since we were especially interested in CSF IgG, patients were selected for a high intrathecal IgG production according to the Reiber formula [IgG) mean 36 %, median 44 %, interquartile range (IQR) 9 to 52 %). From these 27 MS cases, 18 had a recent (<90 days) clinical relapse . Nine patients had received steroids within 90 days, and one patient natalizumab. Two patients had concomitant uveitis, one of whom received methotrexate. We analyzed 21 CSF/serum sample pairs from control donors, including 5 healthy control volunteers (HC) and 16 patients with other, non-inflammatory neurological disease . From the donors described, CSF and serum was analyzed, but some mass spectrometric peaks could not be quantified, so that 2 % of data points were missing. From four additional donors , serum profiles could be obtained.We evaluated 48 paired serum and CSF samples. Thereof, 27 were from patients with definite MS diagnosed according to the 2005 McDonald criteria or a first demyelinating event  highly suggestive of incipient MS .QIgG) is defined as the ratio of the concentrations of IgG in CSF divided by IgG in serum. As a more elaborate method to quantify the fraction of IgG that is produced intrathecally (IFIgG), we applied this formula  across all comparisons (sugar residues and IgG subclasses) as family-wise error rate (Bonferroni correction). When depicting glycosylation as absolute values in CSF and serum separately (=2 compartments), this was also taken into account, resulting in a higher correcting factor for absolute CSF and serum values (4 sugars × 2 IgG subclasses × 2 compartments = 16), compared to CSF/serum ratios (4 sugars × 2 IgG subclasses × only 1 ratio of both compartments = 8). Boxplots were plotted with default whiskers from R (range up to 1.5× IQR below/above first/third quartile).Comparisons between two groups were calculated by Mann-Whitney p values were adjusted for multiple testing across all possible comparisons of glycoforms and (para-)clinical observations including galactosylation, sialylation, bisecting GlcNAc, afucosylation, age at LP, disease duration, time from last relapse, CSF cell count, Qalb, QIgG, IFIgG, EDSS) as false discovery rates [locally weighted regression and smoothing scatterplot (LOWESS) lines [x-axis, data points were jittered horizontally within each category to avoid them obscuring each other. Colors were chosen for best contrast also for colorblind people. We noted that MS cases differed from controls regarding IgG glycosylation in CSF, but not in serum, and normalized CSF glycosylation data as CSF/serum ratio. Using CSF/serum ratios instead of absolute CSF glycosylation values reduced noise  by about 60 %, but did not affect the direction of differences in between the groups  lines . When plps Figs.  and 4. Tprincomp from the stats package in R. Principal component analysis axis scales for score and loading were plotted according to the biplot standard.Principal component analysis was performed by the function We quantified IgG1 Fc glycosylation in CSF and serum. A scheme of the attached oligosaccharides, along with frequencies of the respective variable sugar residues in control CSF IgG1, is shown in Fig. We noted differences between CSF and serum IgG1 glycosylation Fig. . In CSF absolute level of glycosylation differed between CSF and serum, we observed a positive correlation between identical IgG1 glycoforms in serum vs. CSF  than for the MS group , consistent with an intrathecal IgG production in addition to plasma-derived IgG in the CSF from MS cases. Correlations between serum and CSF IgG2 were less pronounced as compared to IgG1 but present especially for IgG2 galactosylation both in MS and controls .While the CSF Fig. . This coBisecting GlcNAc of IgG1 was increased in CSF from MS cases. This was evident by the CSF/serum ratio  did not reach statistical significance . The CSF/serum ratio of afucosylated IgG1 glycoforms was decreased in MS compared to control donors  and present as a trend after correction for multiple testing for galactosylation and bisecting GlcNAc.MS-related changes in Fc glycosylation were more pronounced in those patients with higher cell counts Fig. . The corQIgG and the intrathecally produced IgG fraction (IFIgG) according to the Reiber formula [QIgG; ϱ = −0.65, p < 0.001, padj = 0.004; IFIgG; ϱ = −0.61, p < 0.001, padj = 0.012). This is consistent with an intrathecal production of less afucosylated IgG, as might be inferred also from the group comparisons  and same group (MS cases or controls), we noted that by far the strongest positive correlation existed between sialylation and galactosylation .Despite these absolute differences, IgG1 and IgG2 glycosylation was related. In CSF, this correlation of respective IgG2 and IgG1 glycoforms was much weaker and only present as a trend for most glycosylation features, especially in the MS group .In This study shows four main results. First, IgG1 glycosylation differed between CSF and serum, even in the control group without intrathecal IgG synthesis. Second, in MS patients vs. controls, the IgG1 glycosylation pattern was altered in CSF, but not in serum. Third, alterations of glycosylation occurred especially shortly after a relapse. Fourth, glycosylation patterns in CSF from MS cases correlated with the degree of intrathecal IgG1 synthesis and CSF cell counts.IgG glycosylation regulates Fc effector functions. A pro-inflammatory pattern consists of elevated bisecting GlcNAc but reduced galactosylation , as we oopposite effect, namely reduction of complement activation by afucosylation in the case of a therapeutic CD20 depleting antibody [For the decrease of afucosylation, as observed in CSF IgG1 from MS patients, the interpretation is more complex; afucosylation has initially been regarded as pro-inflammatory by enhancing ADCC via FcγRIIIa . A recenantibody . Thus, tantibody , and in antibody . Neverthantibody . Therefoantibody .inter-group-level, but were linked to signs of inflammation also within the MS group, especially that the alteration in afucosylation  was more pronounced in those patients with stronger signs of intrathecal inflammation .Of note, MS-related changes in CSF IgG glycosylation where not only evident on the When all four glycosylation features were combined by principal component analysis, separation of the groups was of similar discriminatory power as established markers such as CSF cell count or the intrathecal IgG fraction. However, there is no evidence that this inflammatory pattern of glycosylation is MS specific, but might rather be associated with the degree of inflammation. Using the glycopatterns and type of analysis we describe here, future studies can now apply these methods to address a number of obvious issues, such as the predictive value in addition to oligoclonal bands, comparison of autoimmune and infectious CNS diseases, alterations during aging, and effects of immunotherapy.IgG). Third, the strong correlation of CSF and serum IgG glycosylation in control donors was much weaker in the MS group, where intrathecally produced IgG occurs in addition to circulation-derived IgG. Fourth, the group differences between controls and MS patients were greater for IgG1 than for IgG2, and intrathecally produced IgG is mainly IgG1 [We conclude from our data that it is the intrathecal production of IgG that results in an altered glycosylation pattern for the following reasons. First, MS-related glycosylation abnormalities were present in CSF, but not in blood. Second, the degree of these abnormalities (esp. afucosylation) in the CSF from MS patients correlated with the fraction of IgG that was actually produced intrathecally , the kinetics were compatible with an inflammatory glycosylation pattern building up at the time of a relapse. It is possible that incorporating data on subclinical MRI activity and lesion localization would result in even better correlation of glycosylation patterns and disease features, but frequent MRI data were not available.IgG glycosylation features were largely independent of each other within the same group and compartment, except in the case of sialylation and galactosylation, which highly correlated with each other. This seems plausible, given the subsequent addition of terminal sialic acid on top of galactose Fig. . HoweverAn unexpected finding in this study was that IgG glycosylation in the CSF is distinct from serum even in the absence of inflammation and intrathecal IgG production. Although these differences were small, they were significant also after adjustment for multiple testing. Potential explanations include that (1) IgG transport into or (2) half-life within the CSF compartment depends on glycosylation, or that (3) IgG glycosylation is modified in the CSF or serum.Galactosylation was reduced in control CSF and could thus favor IgG effector functions also in healthy subjects, but it is likely that a further reduction of galactosylation, as observed in MS Fig. , as wellSince clinical samples were collected at different sites, we cannot completely rule out any site bias. However, in order to minimize any pre-analytical issues, samples were collected following the same protocol, centrifuged and frozen immediately, shipped on dry ice, and analyzed by mass spectrometry altogether as detailed in the Methods section. Analyzing the CSF/serum ratios of the MS vs. the control group separately for site 1 and site 2, we observed similar changes also within the samples of site 1 or 2 for the glycofeatures with significant regulation, arguing against a major site bias.The CNS compartment as well as the inflammatory milieu in MS affect IgG1 Fc glycosylation. In MS, the altered CSF IgG1 glycosylation pattern has pro-inflammatory features and is linked to intrathecal IgG synthesis (see also Fig.


During my 40 years of involvement with hand surgery, I have witnessed the following major breakthroughs:- Tendon surgery and tendon program- Development of artificial finger joints- Microsurgery- Various fixation systems for fractures- Wrist arthroscopyCertain breakthroughs are appearing in the horizon:- 3-D printing- Tissue engineering- Genetic engineering- Intra-uterine surgeryAt organizational level, the following are also important:- Team approach for rehabilitation- IFSSH, APFSSH, national societiesHowever, all these technical and scientific advances should be guided by future breakthroughs in the arts and humanities of medicine – the love and care for our patients.


The neurobiological underpinnings of avolition in schizophrenia remain unclear. Most brain imaging research has focused on reward prediction deficit and on ventral striatum dysfunction, but findings are not consistent. In the light of accumulating evidence that both ventral striatum and dorsal caudate play a key role in motivation, we investigated ventral striatum and dorsal caudate activation during processing of reward or loss in patients with schizophrenia.We used functional magnetic resonance imaging to study brain activation during a Monetary Incentive Delay task in patients with schizophrenia, treated with second-generation antipsychotics only, and in healthy controls (HC). We also assessed the relationships of ventral striatum and dorsal caudate activation with measures of hedonic experience and motivation.The whole patient group had lower motivation but comparable hedonic experience and striatal activation than HC. Patients with high avolition scores showed lower dorsal caudate activation than both HC and patients with low avolition scores. A lower dorsal caudate activation was also observed in patients with deficit schizophrenia compared to HC and patients with non-deficit schizophrenia. Dorsal caudate activity during reward anticipation was significantly associated with avolition, but not with anhedonia in the patient group.These findings suggest that avolition in schizophrenia is linked to dorsal caudate hypoactivation. Additional eligibility criteria were: no family history of mental illness or psychiatric hospitalization; no past history of head trauma with loss of consciousness; no lifetime history of substance abuse or dependence (except for smoking); not on prescribed medications that might affect CNS functions.The study was approved by the University Ethics Committee, and all participants signed a written informed consent.All subjects were interviewed to record socio-demographic variables, including age, education and socioeconomic status. An estimate of full-scale IQ was obtained using the revised version of the Wechsler Adult Intelligence Scale.et al.et al.et al.  and several other studies carried out in patients with schizophrenia .et al.The task difficulty was personalized during a practice session prior to scanning according to the original MID task design  to avoid the potential effects of nicotine withdrawal.2), providing 32 interleaved images per volume, parallel to the AC–PC line and covering the whole brain. Each fMRI series consisted of 368 images, the first four of which were discarded to allow the scanner to reach a steady state.Structural and functional images were acquired on a 3.0-T scanner , equipped with a standard radio-frequency head coil. Head movements were restricted using foam cushions. Structural images were acquired via a high resolution, T1-weighted 3D MPRAGE sequence . T2*-weighted functional images covering the whole brain were acquired using a GRE-EPI sequence depicting an event-related blood oxygen-level dependent (BOLD ) signal  was used. Preprocessed images were corrected for differences in slice-time acquisition, realigned to the mean volume, and spatially normalized to the standard template of the Montreal Neurological Institute (MNI). The spatially normalized data were smoothed with an isotropic Gaussian filter  to compensate for normal variation across subjects.For image preprocessing and GLM analysis, the SPM8 software package  corrections for multiple comparisons were performed on all results, except for the ventral striatum and dorsal caudate, for which small volume statistics were applied, for which a family-wise error (FWE) peak correction was deemed appropriate, as these are considered appropriate for small structures, where a relatively small number of voxels per cluster is to be expected  who investigated ventral striatal activation in patients with schizophrenia and its associations with psychopathology. Online Supplementary Fig. S2 illustrates the VOI boundaries.Whole brain random-effects statistical maps were thresholded at .et al. b. For th .For correlations, measures of brain activation  for each contrast were extracted from the above VOIs as well as from VOIs of key cortical regions connected with ventral striatum and dorsal caudate, namely OFC and DLPFC, using the MARSBAR toolbox  was used to test group differences on continuous variables.Group differences in sex distribution were assessed by the Pearson's χZ test, if at least one correlation with the scores for motivation/avolition or anhedonia was significant. Since multiple correlations were computed between VOIs and the above scores, a Bonferroni correction for multiple test was applied and will be reported; however, since Bonferroni correction can bias results toward type-2 statistical error, particularly for a relatively small sample size, and given our a priori hypothesis of the direction of the correlations , only Bonferroni correction for one-tailed alpha = 0.05 will be reported (Bonferroni-corrected p = 0.008). Since antipsychotic treatment, depression severity and positive symptoms might influence striatal activity, anhedonia and motivation, correlations between these variables were examined using partial correlation analyses controlling for daily antipsychotic dose, positive symptoms scores and depression scores.Relationships of measures of brain activation in the ventral striatum, dorsal caudate, OFC and DLPFC with anhedonia, motivation and avolition were examined in patients using Pearson's correlation coefficients. Intercorrelations between anhedonia and avolition/motivation measures were examined to verify whether these were distinct constructs or measured partially overlapping constructs. Differences in the correlation pattern between striatal VOIs  or connected VOIs  were examined using Steiger's Twenty-eight patients with schizophrenia and 22 HC completed the study. There was no group difference in sex, age, parental education, socioeconomic status or proportion of habitual cigarette smokers . No currF4,84 = 8.12, p < 0.00001), due to the fact that they were faster for small reward (p < 0.002) and large reward (p < 0.00001) than for neutral cues. Patients did not show a significant effect of the cue . There was no main effect of diagnosis  and no cue × diagnosis interaction .Details of MID task performance are shown in online Supplementary Table S1. For reaction time, a main effect of the cue was found in controls only  and the same was true for anhedonia and motivation . Furthermore, avolition and motivation scores were not correlated with TEPS score for anticipatory pleasure . These results indicated that the constructs of anhedonia and avolition are at least partially independent and that avolition and motivation do not derive from a deficit in the ability to anticipate pleasure. On the other hand, avolition was significantly correlated to motivation in the real world . This latter correlation indicates that avolition and motivation are intercorrelated constructs.Correlations between anhedonia and avolition were modest and not statistically significant  was found with real-life motivation , and a negative correlation  was observed with avolition . The correlation between the left dorsal caudate and avolition remained significant after controlling for the effects of PANSS depression and positive symptoms factors, as well as chlorpromazine equivalent dose .For the ventral striatum activity during anticipation, no significant correlation with anhedonia, motivation and avolition was found in patients. For the dorsal caudate activity during anticipation for the contrast reward v. neutral were tested using the Steiger test for dependent correlations and found to be statistically significant Differences in the association of avolition/motivation with activity of the dorsal caudate or ventral striatum for the contrast reward v. neutral, no significant correlation was found with anhedonia, motivation and avolition.For the dorsal caudate activity during anticipation for the contrast loss No significant correlation was observed for the feedback analyses.v. low avolition scores on ventral striatum and dorsal caudate activation and on demographic, psychometric and clinical variables.A median split analysis was carried out to compare patients with high F1,26 = 0.71, p = 0.41), education  and IQ . The high-avolition group, in comparison with the low-avolition group, was receiving a higher chlorpromazine equivalent dose of antipsychotic . The chlorpromazine equivalent dose was entered as covariate when the two subgroups were compared on striatal activity. No difference was found between the two subgroups with respect to the scores on PANSS depression and positive symptom factors, TEPS and PAS. Real-life motivation was significantly lower in high- v. low-avolition patients .High- and low-avolition subgroups did not differ on age  with group  as between factor and education and IQ as covariates did not show any significant main effect of group ; the same analysis on the left dorsal caudate activity yielded a significant group effect  and Bonferroni post-hoc test revealed a significantly reduced activity only in the high-avolition subgroup v. HC .ANCOVA on the ventral striatum activity  did not show any significant main effect of group ; while the same analysis on the left dorsal caudate activity yielded a significant group effect , due to lower activity in the high-avolition subgroup .Comparison between the high- and low-avolition subgroups, with chlorpromazine equivalent dose as covariate, on the ventral striatum activity  and 17 as having NDS (only 3/17 were in the high-avolition group).F1,26 = 9.96, p < 0.004), higher PAS  scores and, as expected, higher avolition  and lower motivation  scores. IQ and PAS scores were entered as covariate when comparing the two subgroups on striatal activity.DS and NDS subgroups did not differ on age, education, PANSS depression and positive symptom factors, TEPS and chlorpromazine equivalent dose of antipsychotic. Compared with the NDS subgroup, DS patients had significantly lower IQ  with group  as between factor and education and IQ as covariates did not show any significant main effect of group . ANCOVA on the left dorsal caudate activity for the same contrast yielded a significant group effect ; Bonferroni post-hoc test revealed a significantly reduced activity only in the DS subgroup v. HC .ANCOVA on the ventral striatum activity  ; while the same analysis on the left dorsal caudate activity (for the same contrast and with the same covariates) yielded a significant group effect , due to lower activity in the DS subgroup .Comparison between the two subgroups of patients on the ventral striatum activity for the same contrast, with PAS scores and IQ as covariates, did not show a significant group effect (a) the activation of these two regions during processing of reward or loss in persons with schizophrenia living in the community and stabilized on treatment with SGAs only, and (b) the relationships of ventral striatum and dorsal caudate activation with hedonic experience and motivation.The present study aimed to assess the role of ventral striatum and dorsal caudate in motivation deficits of individuals with schizophrenia. To this aim, we investigated: ; and task difficulty was personalized for each participant, so that they could succeed on at least 66% of the trials.Treatment of patients with antipsychotic drugs might represent a further limitation, as in the patient group the lack of dorsal striatum activation during reward anticipation might be attributed to the treatment with SGAs. However, antipsychotics would be expected to dampen the response to reward in all sections of the striatum, since all of them receive dopamine innervation, while we observed in our patients a significant activation of the ventral striatum during reward anticipation, a pattern similar to that observed in healthy subjects. These results argue against a role of SGAs in our findings.Another limitation is the assessment of extrapyramidal symptoms only during the routine neurological examination of our patients, without using a standardized instrument. Extrapyramidal symptoms might cause secondary negative symptoms which might confound results concerning avolition. However, we think it is unlikely that extrapyramidal symptoms affected our findings concerning avolition. First, subjects with primary and persistent negative symptoms (deficit schizophrenia), identified using the SDS, were the majority of the subjects with high avolition and, by definition, the influence of extrapyramidal symptoms on avolition in this group had to be excluded. Second, our subjects had a preserved ventral striatal response to reward, that would be unlikely in the presence of extrapyramidal side-effects.Finally, to measure real-life motivation, we used an index derived from the QLS scale, and healthy subjects showed a ceiling effect on this instrument, preventing the exploration of correlations between motivation and striatal activation in this group.In conclusion, our findings support the notion that hedonic and motivational aspects of reward are subtended by different subdivisions of the striatum; that avolition in schizophrenia emerges independently of in-the-moment ability to experience pleasure, and that it is not linked to a ventral striatum dysfunction (at least in patients treated with SGAs) but to the hypoactivation of the dorsal caudate.et al.et al.et al.Our finding of a dorsal striatum hypoactivation in patients with schizophrenia is of interest also in the light of the recently documented role of the human dorsal striatum in complex social tasks, such as social interactions in situations requiring cooperation (Rilling


Spontaneous intracranial hypotension (SIH) is a rare but increasingly diagnosed condition. Low cerebrospinal fluid (CSF) pressure due to a dural leakage causes orthostatic headache. The nature, etiology, and location of the CSF leak itself are currently unknown and are thought to be spontaneous or idiopathic in most cases.We present a case series with proven CSF leaks in which a systematic and meticulous search revealed dura-perforating micro-spurs as a cause of SIH.A consecutive series of patients with symptoms of intracranial hypotension were evaluated for a systematic diagnostic work-up from February to August 2013. We performed a spine focused, stepwise escalating imaging set. If a CSF leak could be restricted, we performed microsurgical exploration under intraoperative electrophysiological monitoring of the presumed site of the CSF-leak.We identified 6 patients with SIH and imaging signs of spinal dural perforation. In 5 cases a calcified micro-spur extruding out of the disc space was identified perforated the dura and arachnoid and was the cause of CSF leak in all 5 cases. In one case a broad osteophyte had ripped the dura. All 5 micro spurs and one osteophyte were micro-surgically removed, the dura was sealed, and the CSF leak ceased immediately.The etiology of the CSF leak in SIH remains obscure. Here we present 6 patients in which a systematic spinal work-up, including microsurgical exploration, revealed dura perforating micro spurs. The dura-perforating micro spurs were a frequent, definitive, and readily treatable cause of SIH.No conflict of interest.


ObjectRetrosigmoid (RS) and translabyrinthine (TL) surgery remain essential treatment approaches for symptomatic or enlarging acoustic neuromas (ANs). We compared nationwide complication rates and payments, independent of tumor characteristics, for these two strategies.MethodsWe identified 346 and 130 patients who underwent RS and TL approaches, respectively, for AN resection in the 2010-2012 MarketScan database, which characterizes primarily privately-insured patients from multiple institutions nationwide.ResultsAlthough we found no difference in 30-day general neurological or neurosurgical complication rates, in TL procedures there was a decreased risk for postoperative cranial nerve (CN) VII injury , dysphagia , and dysrhythmia . Overall, there was no difference in surgical repair rates of CSF leak; however, intraoperative fat grafting was significantly higher in TL approaches . In patients receiving grafts, there was a trend towards a higher repair rate after RS approach, while in those without grafts, there was a trend towards a higher repair rate after TL approach. Median total payments were $16,856 higher after RS approaches , without differences in physician or 90-day postoperative payments.Conclusions Using a nationwide longitudinal database, we observed that the TL, compared to RS, approach for AN resection experienced lower risks of CN VII injury, dysphagia, and dysrhythmia. There was no significant difference in CSF leak repair rates. The payments for RS procedures exceed payments for TL procedures by approximately $17,000. Data from additional years and non-private sources will further clarify these trends. Surgical intervention remains a viable treatment option for symptomatic or progressively enlarging acoustic neuromas (AN) . The mosIn general, the RS approach is considered more versatile, enabling removal of tumors largely independent of size. In contrast with a TL approach, an RS resection can offer hearing preservation . The RS Although the advantages and disadvantages of different approaches for AN resection have been extensively reported on, a large-scale study to enable their accurate comparison is still greatly needed. The absence of a unified, consistent reporting of variables may contribute to variations in procedure outcomes. For example, in their meta-analysis, Ansari, et al. reported that the RS approach was the most versatile for facial nerve preservation, but it also led to a higher risk of postoperative pain and cerebrospinal fluid (CSF) fistula relative to the middle fossa or TL approach . MeanwhiWe performed a retrospective, longitudinal analysis of RS and TL approaches in procedures treating acoustic neuromas from a national database from years 2010–2012. Inpatient and outpatient data were obtained from the Thomson Reuters MarketScan Commercial Claims and Encounters and Medicare Supplemental and Coordination of Benefits databases, administered by Truven Health Analytics. The MarketScan dataset includes data from over 100 payers and includes inpatient, outpatient, and pharmacy services from a range of large employers, health plans, governmental organizations, and public organizations.We identified patients undergoing RS procedures with Common Procedural Terminology (CPT) code 61520 and TL procedures with CPT code 61526. A concurrent International Classification of Diseases (ICD-9) diagnosis code of 225.1 was required to indicate the presence of an acoustic neuroma. Comorbidities were assessed using the Deyo-Charlson and Elixhauser ICD-9 code groupings . SupplemWe identified surgeons using anonymized payer-specific physician identification numbers within inpatient records from the MarketScan data set. In some cases, a physician may have multiple identification numbers not reconciled among payers; however, due to the anonymization, we considered each physician number as unique. We then calculated the annual volume per individual surgeon identifier.Tests of significance were performed using two-tailed tests. Categorical variables were analyzed using Fisher’s Exact test; continuous variables were analyzed using Student’s t-test. Due to the examination of a range of complications, significant results (P < 0.05) were adjusted with the Holm-Bonferroni method to determine whether the hypothesis could be rejected. Data preparation and analysis were performed using SAS software .Within the MarketScan database during the studied period between 2010 and 2012, there were 346 and 130 patients who underwent an RS and TL approach for acoustic neuroma resection, respectively. There was no significant difference between the RS and TL groups for any of the patient factors, comorbidities, or hospitalization characteristics examined, shown in Table There was no significant difference in the incidence of overall general neurological or neurosurgical complications between the two procedures within the 30-day postoperative period, as shown in Table While there was no difference in the rate of lumbar drain placement  or surgical repair of CSF leak , the use of a fat graft during surgical repair of a CSF leak was significantly higher in the TL approach , shown in Table In patients with grafts, the need for a CSF leak repair trended towards being higher in the RS group . However, in those without fat grafts, it was higher in the TL group . Of note, we also found that patients who underwent the TL approach and received fat grafting were less likely to develop CSF leak requiring surgical intervention . In contrast, there was a trend towards increased risk of CSF leak requiring surgical intervention with fat grafting in patients who received an RS approach .The RS approach resulted in higher median total payments  and hospital payments . There was no significant difference between the two procedures for median physician payments  or aggregate 90-day postoperative payments .  There was no difference in the length of hospital stay, follow-up, or discharge home. These findings are shown in Table Surgeons who are recorded in the MarketScan database as performing at least two AN procedures annually had a decreased incidence of CN VII injury  and postoperative dysphagia , as well as a trend towards decreased need for repair of CSF leak , compared to surgeons performing fewer than two procedures per year. These results are shown in Table This national sample of patients undergoing surgical intervention for acoustic neuromas between 2010 and 2012 compared the complications and costs between these two common approaches. We observed that an RS approach, relative to a TL one, was associated with an increased rate of postoperative facial nerve injury (18.7% vs 2.6%) and postoperative dysphagia (8.9% vs 2.6%). Although the frequency of CSF leak was equivalent between approaches, the choice of intraoperative fat grafting reflected each procedure’s respective needs for adequate dural repair . Total hFollow-up analyses did not indicate that variations in the length of stay or lumbar drain placement contributed to the difference in costs between RS and TL approaches. However, the diverging postoperative complication rates, particularly in relationship to provider volume, may play a role. Prior work has reported that high volume centers achieve a lower average cost of hospitalization -19. Low The rate of facial nerve dysfunction associated with AN surgery is an important clinical outcome. Affecting one out of five RS patients in our analysis, it was the most prominent individual complication in the early postoperative phase. The TL approach is traditionally thought to provide a more anatomically accommodating view of the facial nerve and, therefore, can limit facial nerve injury . ExistinMeanwhile, other studies emphasizing additional parameters or with longer follow-up have reported varying complication rates. For example, Ansari, et al. reported that at 23.3 months median follow-up, there was no difference between surgical approaches when treating smaller tumors and that an RS approach can actually have better facial nerve preservation rates when treating larger tumors . Since fAlthough less common than facial nerve dysfunction, postoperative injury to the neighboring vagus nerve, presenting as dysphagia, is another important clinical outcome for patients and providers. The complication is again largely limited to the early postoperative phase, with follow-up surveys indicating that 71% recover from their vagal palsies. Those with palsies were no more likely than those who never presented with palsies to have long-term difficulties related to their voice or swallowing . NeverthInterestingly, the rate of CSF leak is comparable between surgical approaches. Although there remains no consensus on the best graft protocol for preventing postoperative CSF leaks, there is encouraging data showing that fat grafting can reduce the rate of a dural leak to 0-7.4% -37. WithFinally, we observed significant variation in inpatient financial impact with AN resections. The difference in median total payment exceeded $15,000 between the two approaches, and this margin is likely to play an increasingly larger role in clinical decision-making as the cost of AN surgery continues to rise -39. SincOur study has several limitations, which should be noted. The MarketScan dataset does not characterize tumor size or location , which frequently guides the surgical approach. Our analysis is intended as a broad overview of the comparative outcomes of the procedures independent of preoperative evaluation and planning. Secondly, the MarketScan database is limited in assessing surgical volume or providers, as it only captures procedures whose payment is provided by insurers within the database. Our assessment of volume is, therefore, more reflective of relative, and not absolute, volume differences. We are also not able to differentiate between academic and private practices, which frequently vary in case distribution and complexity. Outcome differences may be attributable to surgeons' experience with acoustic neuroma management and the approaches for resection. In our analysis of CSF leaks, it is important to note we consistently discussed the rates of CSF leak requiring surgical intervention. We are unable to report the objective rates, as the ICD-9 codes that may provide that information is unreliable. Furthermore, because cranial nerve palsies have been shown to resolve and hospital costs are most prominent during the initial hospitalizations, we believe our data appropriately focuses on the symptomatically and resource-intensive period relevant for accurate quality assessment.Our report, which is accompanied by the limitations of all retrospective studies, provides an updated, large-scale survey of current AN surgical care. These results can serve to inform future evaluations of both community- and academic-based practices.From a nationwide, administrative database spanning 2010-2012, AN resection by TL approach was found to have a lower rate of CN VII injury, dysphagia, and dysrhythmia compared to that by RS approach. Although there was no significant difference in CSF leak repair rates, the rate for tissue grafting differed by approach. Surgeons performing multiple AN resections had lower rates of CN VII dysfunction and lower CSF leak repair rates, independent of the procedure. The mean payment for RS procedures was significantly greater than that for TL procedures; however, there was no difference in post-hospitalization costs.


The authors describe a case of cerebrospinal fluid rhinorrhea due to a congenital defect in cribriform plate, an anomaly that has not been described hitherto. It was successfully treated surgically.  Cerebrospinal fluid (CSF) rhinorrhea is commonly reported with traumatic brain Injury, ruptured anterior encephalocele, brain tumors; idiopathic spontaneous CSF rhinorrhea is also known [1]. Spontaneous anterior fossa CSF rhinorrhoea and congenital perilymphatic fistula has been reported [2]. We report a neonate with normal intra-cranial pressure CSF rhinorrhea due to congenital defect in cribriform plate, and briefly describe the diagnostic difficulties and surgical management.A 27-day-oldneonate was brought with complaint of continuous watery discharge from left nostril from 2nd day of birth. He was a full term baby delivered at 36 weeks by LSCS with 9/10 APGAR score at 1min. There was no history of birth trauma, fever, seizures and other symptoms of raised intra-cranial tension or intracranial infection. The neonate was diagnosed as viral rhinitis by the pediatricians elsewhere and had been already administered 4 courses of anti-histaminics and antibiotics before he was brought to us in a dehydrated state. Examination of the baby was unremarkable without any external cranial trauma, any visible encephalocoele, normal inter-pupillary distance . Beta-transferrin was positive confirming that the fluid was CSF. Cranial MRI revealed normal brain without any evidence of tumor or encephalocoele, but a continuous CSF flow in left nostril . 3D MR reconstruction of skull base revealed defect in the left cribriform plate . Crista galli was deviated to the right. In view of the fact that there was no expertise to perform endoscopic trans-nasal repair and small endoscopes were unavailable in such a small baby, was contemplated but the senior author (GRP) decided to repair the defect through trans-frontal trans-dural approach. The defect was exposed by bi-temporal trans-frontal craniotomy. The skin flaps were raised up to the eyebrows. The dura was exposed by raising a laterally placed osteoblastic flap . It was opened with two dural tacker sutures in situ. The defect was exposed  and was plugged with abdominal wall fat, neuro-surgicel and a free peri-cranial graft (fig. 2c) Dura was closed continuous inter-locking sutures . The osteoblastic flap was restored followed by closure of skin. Instantaneous cessation of CSF leak was observed at the end of surgery . The post-operative period was uneventful. Follow up MRI after 9 months showed plug in place. . On 7 years follow-up, the child is thriving and growing well. Unilateral watery nasal discharge in an otherwise normal child should always be suspected to be CSF rhinorrhea. Unilateral, watery, clear rhinorrhea is extremely rare in viral rhinitis [3]. Unilateral nasal discharge in older children is a symptom qua-nun of foreign body nose. A reservoir test simply denotes the ability to collect the watery discharge from nose into a reservoir. This bedside test is not positive in other causes of nasal discharge. Although biochemical values had suggested CSF, beta-transferrin estimation, which is not universally available, is diagnostic of CSF [4]. MRI would clinch the diagnosis. Although a trial of conservative medical treatment is indicated in other causes of CSF rhinorrhea/ ottorrhea, CSF rhinorrhea with an anatomical cribriform defect will obviously not respond to conservative treatment and the child will be put to risk of meningitis. CSF rhinorrhea can be approached either trans-nasally, trans-sphenoidally or trans-frontally . The open surgical approach has been already justified above.Source of Support: NilConflict of Interest: Nil


High serum Retinol Binding Protein 4 (RBP4) levels were associated with insulin-resistant states in humans. To determine which fat compartments are associated with elevated RBP4 levels in humans, we measured serum RBP4 and hepatic fat content (HFC), visceral (VFA) and subcutaneous abdominal fat area (SFA) in 106 subjects with non-alcoholic fatty liver disease (NAFLD) without known diabetes.106 patients with NAFLD  were enrolled. Subjects with known diabetes, chronic virus hepatitis, and those with alcohol consumption ≥30 g/d in man and ≥20 g/d in woman were excluded. Anthropometrics and laboratory tests, including lipid profile, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and γ-glutamyltransferase (γ-GT) were conducted. HFC, VFA and SFA were determined by CT scan. Serum RBP4 was detected by an enzyme immunoassay kit and validated by quantitative Western blotting.p < 0.001), but positively with waist-to-hip ratio (WHR) , triglyceride , VFA , systolic blood pressure , diastolic blood pressure , γ-GT , waist circumference . Differently, serum RBP4 levels were not associated with HFC , SFA, age, BMI, total cholesterol, low-density lipoprotein cholesterol (LDL-c), ALT or AST . Multiple linear regression analysis revealed that RBP4 correlated independently with VFA  and HDL-c  in all subjects, HDL-c  in men, VFA/SFA in women , not with HFC. However, serum RBP4 was positively correlated with HFC when HFC below 6.34% .Circulating RBP4 was negatively associated with high-density lipoprotein cholesterol (HDL-c) (r = −0.392, RBP4 could be a marker of abdominal obesity, however, the role of RBP4 in the pathogenesis of NAFLD is not sufficiently elucidated. Retinol binding protein 4(RBP4) is a 21 kDa secreted protein that is the sole specific transport protein for vitamin A (retinol) in blood ,4. LiverStudies in mice suggest that elevated serum RBP4 could play a causal role in insulin resistance. Increasing serum RBP4 concentrations causes insulin resistance, by transgenic overexpression or by injection of purified RBP4 protein into wild-type mice . ConversAbdominal obesity is one of the main characteristics of the insulin resistance. Some studies have observed the relationship between RBP4 and abdominal obesity and found that serum RBP4 concentrations in humans correlate highly with waist-to-hip circumference ratio , waist cSo, we speculate that nonalcoholic fatty liver disease (NAFLD), a typical case of visceral obesity, is associate more closely with RBP4. However, research conclusions about the relationship of RBP4 and hepatic fat content are controversial. Some researches indicated that RBP4 is elevated in non-diabetic NAFLD patients , or diabThus, the aim of this paper was to investigate the association between serum RBP4 levels and HFC, visceral (VFA) and subcutaneous abdominal fat area (SFA), as well as other metabolic parameters in Chinese subjects with nonalcoholic fatty liver disease without known diabetes.HFC of all the subjects determined by CT scan varied from 0.56% to 24.7% with a mean and standardized deviation of 7.9% and 6.9%, respectively. By dividing the distribution of HFC into quartile, we found that there were more women subjects than men subjects in groups with lower HFC Q1), but in groups with higher HFC (from Q2 to Q4) there were more men subjects . The four groups did not differ in some metabolic parameters, including age, total cholesterol (TC), low-density lipoprotein cholesterol (LDL-c), alkaline phosphatase (ALP). No significant differ were found in some metabolic parameters in the four groups, including age, total cholesterol (TC),low-density lipoprotein cholesterol (LDL-c), alkaline phosphatase (ALP).Accompanied with the increase of HFC, BMI, waist circumference, WHR, systolic blood Pressure (SBP), diastolic blood pressure (DBP), triglyceride (TG), alanine aminotransferase (ALT), aspartate aminotransferase (AST), γ-glutamyltransferase (γ-GT), VFA and SFA elevated gradually, high-density lipoprotein cholesterol (HDL-c)fell gradually  together with HDL-c  were independent variables significantly associated with serum RBP4. In women, VFA/SFA  was independently associated with serum RBP4, while in men, HDL-c  was variable significantly associated with serum RBP4 ; However, the association between HFC and RBP4 no longer existed when HFC within Q3 to Q4 (more than 6.34%) .In light of the data mentioned above, we analyzed the association between serum RBP4 concentration and HFC in subjects within the first two quartiles of HFC and all subjects, respectively Figure . When HFAlso, a stepwise multiple linear regression analysis was performed in subjects whose HFC were less than 6.34%, using HFC as a dependent variable, RBP4 as an independent variable together with weight, BMI and waist circumference, which are significantly associated with hepatic fat content. And the results showed that RBP4 was significantly associated with HFC . And we found the liver fat content determined by CT was undervalued, hence, some patients might not have NAFLD and still were included in this study due to the diagnostic inferiority of using a CT scan. So, the results come from this study could just be considered as a clue, and the accurate relationships of RBP4 and fat compartments needs further verification by large scale studies and precise method of assessing hepatic fat content, such as liver biopsy or 1H MRS. Secondly, our study is a cross-sectional study, which indicate that cause and effect relationships between RBP4 and body fat composition can not be precisely determined. Thirdly, RBP4 must be bound to retinol for efficient secretion from hepatocytes, retinol deficiency reduces serum RBP4. We do not determine retinol serum concentration and therefore are not able to calculate the theoretic saturation of RBP4 with retinol, which may affect the relationship of RBP4 concentration and metabolic parameters.There are several limitations in this study. Firstly, CT scan is used to determine the hepatic fat content, and its diagnostic accuracy is inferior to pathological diagnosis by liver biopsy and 1H Magnetic Resonance Spectroscopy  were identified and recruited from the outpatient department of Endocrinology of Zhongshan hospital between Nov. 2005 and Apr. 2006 who were diagnosed as fatty liver by ultrasound. Exclusion criteria were alcohol consumption reported by themselves and their families of more than 30 g/d in the case of men and more than 20 g/d in the case of women. Subjects were also excluded if they had known diabetes, hepatitis B or C or other liver diseases, or if they were receiving hypoglycemic or lipid-regulating  drugs, or other drugs could impact hepatic fat content four weeks before the time of enrollment, such as silybin, ursodeoxycholic acid, polyene, phosphatidylcholine, vitamin E, some Chinese herbs, etc. Other exclusion factors were diseases affect the metabolic state or other circumstance not suitable to participate in this clinical trial. These details were described in our previous article .Clinical data such as age, gender, height, weight were collected. Body mass index (BMI) was calculated as the weight in kilograms divided by the square of the height in meters. Waist circumference was measured at the midpoint between the inferior costal margin and the superior border of the iliac crest on the midaxillary line. Waist–hip ratio (WHR) was calculated as waist circumference divided by hip circumference. Blood pressure was measured three times with 5 minutes intervals each time in the seated position with a mercury sphygmomanometer in the morning, and the average of the three blood pressures was used as the final blood pressure.Data on demographic variables, health status, health behavior, and physical activity were collected using a standardized questionnaire. All participants were required to fast overnight (12 h) before physical examination by trained staff and physicians using standard protocols. Blood samples were drawn after an overnight fast and immediately centrifuged. Serum glucose was measured by the glucose oxidase method. Serum total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL-c), low-density lipoprotein cholesterol (LDL-c), and liver enzyme levels were determined by enzymatic methods with a Hitachi 7600 analyzer . Serum RBP4 (μg/ml) was measured in duplicate by a sandwich ELISA developed in-house, using affinity chromatography purified polyclonal and monoclonal antibodies generated against recombinant human RBP4 protein. The assay system was subsequently cross validated by Western blot analysis. The intra-assay coefficient of variation was 1.8–7.6% and inter-assay was 3.7–8.85% . The stuThe abdominal adipose tissue areas were quantified by CT scans using a Marconi MX8000 machine (Philips) by the same one experienced doctor. With the subject in a supine position, a 10-mm CT slice scan was acquired at the L4 to L5 level to measure the total abdominal and visceral fat area. Visceral fat area was quantified by delineating the intra-abdominal cavity at the internal aspect of the abdominal and oblique muscle walls surrounding the cavity and the posterior aspect of the vertebral body. The subcutaneous fat area (SFA) was calculated by subtracting the visceral fat area from the total abdominal fat area. Hepatic fat content was evaluated semi-quantitatively by CT scanning as previously described .p values <0.05 were considered significant.All statistical analyses were performed using SPSS18.0 . Normally distributed data were expressed as means ± SD. Data that were not normally distributed, as determined using Kolmogorox–Smirnov test, were logarithmically transformed before analysis and expressed as median with range. One-way ANOVA was used for comparisons among groups, and multiple testing was corrected using LSD method  or Games-Howell method . Pearson’s correlations and multiple stepwise regression analysis were used to examine the association of serum RBP4, HFC, VFA, SFA and other parameters. In all statistical tests,


CSF outflow through the cribriform plate near the olfactory nerves and the outflow along brain and spinal nerves are together known as peripheral CSF outflow pathway (PCOP). It is still not clear whether the PCOP has pathogenetic relevance. Our previous clinical observations have indicated that CSF may interact with nerves along the PCOP and in this article we present our finding of CSF outflow demonstrated by myelography in a single patient. We also discuss unexplained experimental pain pathomechanisms against the background of the PCOP hypothesis. We observed that CSF flowed along lumbar nerves in distal direction at a speed of about 10 cm per hour on its way through the tissues, mainly muscles. Total CSF outflow volume at the lumbar site was remarkable. CSF outflow at lumbar nerves was also documented by neuroradiology. It is plausible that CSF signaling serves for interaction with nerves along the PCOP, which could explain previously unknown pathomechanisms in pain generation. Experimental findings of tactile pain hypersensitivity within lumbosacral pain pathways could be explained by releasing of molecules, microparticles, or exosomes into the CSF by mast cells, which then move with CSF outflow along the PCOP and interact with nerves, initiating even retrograde synaptic stripping. It has been traditionally accepted that cerebrospinal fluid (CSF) flows from the subarachnoid spaces (SAS) through the cribriform plate into the cervical lymphatics, and similarly, along all brain nerves and all peripheral nerves into the respective tissues. This CSF outflow can transport antigens from CSF spaces to the extracellular fluid of peripheral tissues, which are then collected into the lymphatic system . CSF outWe performed CSF filtration as experimental therapy in cases of severe therapy resistant depression or schizophrenia, with the hypothesis that LLNI states may be responsible for therapy resistance . During Levy et al  investig1. In general, CSF signaling can be mediated by solutes, especially proteins, cells, exosomes, microvesicles, or microparticles containing powerful signaling molecules ,13.2. CSF flows from the SAS along the PCOP between the epineurium and perineurium, making its way down  the nerves all along its course touching also the neuronal ganglia. It eventually reaches the peripheral tissues at the area where nerves end and CSF outflow winds up to join the extracellular tissue fluid.3. Any pathogenic CSF contents might potentially interact with nerves at all PCOP sites where such interaction is possible, causing pathogenetic consequences at different sites from the SAS to the nerve ends, and even to the neighboring tissues near nerve ends.4. Pathogenic CSF contents may for example be released to the SAS from mast cells within the meninges near the SAS. Molecules and particles released by mast cell degranulation could then reach through CSF outflow all PCOP sites and elicit local effects:a) At peripheral neuronal ganglia the pathogenic CSF contents may easily elicit retrograde synaptic stripping at the spinal cord level and thus contribute to pain hypersensitivity generation  (b) In the experiments by Levy et al , the masc) Unfortunately the CSF outflow patterns and local volumes at various parts of the PCOP appear to be rather speculative. CSF outflow may depend on the posture  and exped) CSF outflow and signaling by solutes along the PCOP may be accompanied by trafficking  of CSF cells along the PCOP. CSF cell trafficking along the PCOP was proven at the cribriform plate, after which it reached the cervical lymphatics ,16,17. IIn a patient undergoing lumbar myelography we observed physiological CSF outflow at lower body nerves, near the injection site . The patThere are still no studies dealing with CSF signaling at the PCOP sites, possibly mediated by various pathogenic CSF contents including solutes, microparticles, microvesicles, and cells. Our case is a rather typical finding, representing on overall CSF outflow velocity in still position of about 10 cm per hour. More conclusive and systematic studies are needed to more precisely determine the CSF outflow volumes and flow velocity along the PCOP, and interpersonal, intrapersonal, and local variability of CSF outflow. It is also important to define possible modulating factors of CSF outflow like age, sex, body mass, or the influence of motor activity. These aspects require systematic studies in animals and in humans.Given the poorly explained mechanisms of pain generation in multiple sclerosis or experimental allergic encephalitis , or in eAnother question of interest was the overall balance between CSF production and CSF absorption outflow, the latter involving the outflow along the PCOP, glymphatic pathway , absorptAlso, it would be interesting to examine the complex CSF flow patterns involving the pulsatile brain ,23 as th


Supplemental Digital Content is available in the text. 64Cu] -d-Phe1,Tyr3-octreotate (64Cu-DOTATATE) to specifically target macrophages via the somatostatin receptor subtype-2 in vivo.A feature of vulnerable atherosclerotic plaques of the carotid artery is high activity and abundance of lesion macrophages. There is consensus that this is of importance for plaque vulnerability, which may lead to clinical events, such as stroke and transient ischemic attack. We used positron emission tomography (PET) and the novel PET ligand [64Cu-DOTATATE uptake in carotid artery plaques before carotid endarterectomy. 64Cu-DOTATATE uptake was significantly higher in symptomatic plaque versus the contralateral carotid artery (P<0.001). Subsequently, a total of 62 plaque segments were assessed for gene expression of selected markers of plaque vulnerability using real-time quantitative polymerase chain reaction. These results were compared with in vivo 64Cu-DOTATATE uptake calculated as the mean standardized uptake value. Univariate analysis of real-time quantitative polymerase chain reaction and PET showed that cluster of differentiation 163 (CD163) and CD68 gene expression correlated significantly but weakly with mean standardized uptake value in scans performed 85 minutes post injection . Subsequent multivariate analysis showed that CD163 correlated independently with 64Cu-DOTATATE uptake (P=0.031) whereas CD68 did not contribute significantly to the final model.Ten patients underwent simultaneous PET/MRI to measure 64Cu-DOTATATE accumulates in atherosclerotic plaques of the carotid artery. CD163 gene expression correlated independently with 64Cu-DOTATATE uptake measured by real-time quantitative polymerase chain reaction in the final multivariate model, indicating that 64Cu-DOTATATE PET is detecting alternatively activated macrophages. This association could potentially improve noninvasive identification and characterization of vulnerable plaques.The novel PET tracer  In addition, many patients without significant carotid stenosis will experience recurrent stroke and could have benefited from endarterectomy.Carotid atherosclerosis is a major risk factor of stroke and transient ischemic attack. Randomized trials have shown that carotid endarterectomy significantly reduces the risk of recurrent stroke in patients with recent symptoms of transient ischemic attack or stroke and at least 50% stenosis of the relevant carotid artery.18F]-fluoro-2-deoxy-D-glucose (FDG). FDG is a glucose analogue that is taken up by high-glucose–using cells, where FDG is trapped by phosphorylation to allow for in vivo tissue glucose metabolism assessment. A large body of evidence has linked FDG uptake to the macrophage contents of high-risk atherosclerotic plaques.4 However, a major drawback of imaging atherosclerosis with FDG-PET is the lack of specificity of the tracer.A quest for new and more sensitive methods for in vivo identification of vulnerable atherosclerotic plaques is needed. Positron emission tomography (PET) for molecular imaging, typically in conjunction with anatomic imaging with computed tomography (CT), is one promising hybrid modality. The molecular tracer of choice has to date primarily been 2--d-Phe1,Tyr3-octreotate (DOTATATE) labeled with a positron emitter. Two retrospective studies in patients with cancer investigated 68Ga-DOTATATE uptake in the coronary arteries6 and the large arteries.7 Both studies indicated an increased tracer uptake in atherosclerotic lesions. Interestingly, it was recently found that focal uptake of 68Ga-DOTATATE and FDG did not colocalize in a significant number of atherosclerotic lesions.7An alternative and potentially more specific target for imaging macrophages in the atherosclerotic plaque is the somatostatin receptor subtype-2, which is highly expressed by macrophages.64Cu as an alternative to 68Ga labeling.864Cu has a shorter positron range and longer half-life potentially improving spatial resolution and allowing for late image acquisition. In addition, we recently introduced the use of hybrid PET/MRI, which allows for more precise identification of the atherosclerotic plaque when compared with PET/CT.9We recently introduced DOTATATE labeled with 64Cu-DOTATATE as an in vivo molecular tracer of atherosclerotic plaque activity. To do so, we compared in vivo tracer uptake with gene expression of molecular markers of macrophage load: cluster of differentiation 68 (CD68) as well as activated macrophages of the M1/M2 subsets: tumor necrosis factor-α (TNF-α) and CD163 in patients undergoing carotid endarterectomy using simultaneous PET/MRI in a prospective clinical trial. In addition, we aimed to establish the optimal time to wait from tracer injection to image acquisition.The aim of this study was for the first time to evaluate Materials and Methods are available in the online-only Data Supplement.Ten patients  with clinical symptoms of stroke or transient ischemic attack were enrolled in the study before clinically scheduled endarterectomy. One (symptomatic) plaque was recovered in toto from each patient and sectioned in 3-mm slices. This yielded different slice numbers per patient according to the physical size of each plaque specimen and came to a total of 10 plaques and 62 slices in all. Detailed patient characteristics can be seen in Table 64Cu-DOTATATE PET emission data. All 10 patients received an early scan and a total of 7 patients completed both an early and a late scan. Evidence of arterial wall thickening was seen both in the internal as well as the external carotid arteries in T1, T2, and PD-weighted imaging. These findings were matched by a pattern of stenotic lumen on time-of-flight–weighted imaging in the affected arteries  for all patients on a slice-by-slice basis. No correlation between MRI assessment of plaque burden and 64Cu-DOTATATE uptake determined by PET was found in univariate analysis (P=0.116).All patients underwent MRI for anatomic evaluation of carotid atherosclerosis simultaneously with collection of 64Cu-DOTATATE PET/MRI scans demonstrating tracer uptake in plaques of the internal carotid artery can be seen in Figures 64Cu-DOTATATE uptake was determined by mean standardized uptake value (SUVmean): early scan  and late scan . SUVmean obtained early was significantly higher than SUVmean obtained later, and a slice-by-slice comparison showed a mean difference of 43.9% . Finally, the SUVmean values were significantly higher in the index lesion compared with the contralateral carotid artery . Figure 64Cu-DOTATATE PET/MRI scans of the internal carotid artery from a single patient at 2 different transaxial levels with heterogeneous and no uptake of 64Cu-DOTATATE, respectively. Overall 64Cu-DOTATATE uptake was heterogeneously distributed throughout the plaques. This finding was corroborated with the recovery of a single plaque that was visualized in toto using a preclinical PET/CT system to show the heterogeneity of tracer distribution ex vivo , CD68 showed , cathepsin K showed , interleukin-18 showed , matrix metalloproteinase-9 showed , and TNF-α showed . Nonparametric testing (Spearman correlation matrix) demonstrated good interplaque correlation and strong statistical significance between the marker of activated macrophages (CD163) and the other molecular markers of plaque vulnerability (Table As for 64Cu-DOTATATE uptake (P<0.001), as well as between CD68 expression and 64Cu-DOTATATE uptake (P=0.015). When entering both CD68 and CD163 into the mixed model, only CD163 remained significant (P=0.031). cathepsin K, interleukin-18, matrix metalloproteinase-9, and TNF-α were not significant when entered individually into the mixed model (Table We used a mixed model to take into account the possible nonindependence of slices from the same patient. When entering 1 molecular marker (univariate), we found a weak but highly significant correlation between CD163 expression and A case study was subjected to immunohistochemical analysis. This confirmed the macrophage presence in the vicinity of and within the lipid core deep in the atheromateous plaque shown by specific CD68 and CD163 staining, which concomitantly colocalized with matrix metalloproteinase-9 expression Figure . Further64Cu-DOTATATE. We found a highly heterogeneous intrapatient uptake within each atherosclerotic plaque. Interestingly, this uptake was significantly associated with biomarkers of macrophage load (CD68) and macrophage activation (CD163) in univariate analysis. This association seemed primarily driven by CD163 positive (CD163+) macrophages. This finding could have special interest because CD163+ macrophages are known to have an important role in plaques with hemorrhagic zones.10 In addition, we found evidence that time from tracer injection to PET acquisition is of paramount importance because tracer accumulation decreases almost 50% from our early scan to our late scan after correction for decay. Finally, we found no correlation between plaque burden and 64Cu-DOTATATE uptake.In this study, we present the first results from simultaneous PET/MRI of human carotid plaques using the novel somatostatin receptor tracer 7 Both previous clinical studies used 68Ga-labeled DOTATATE. A preclinical study demonstrated colocalization of 68Ga-DOTATATE and macrophage-rich plaques by autoradiography in a mouse model.1168Ga has the advantage of being generator-eluted and thus has no need for an onsite cyclotron. However 68Ga has high maximum positron energy of 1.899 MeV witch translate into a high-positron range in water; 8.2 mm (maximum) and 2.9 mm (mean) and thus diminished spatial resolution. This is of vital importance for imaging small objects like carotid plaques. To circumvent this, we recently introduced 64Cu-labeled DOTATATE.864Cu is a low-energy positron emitter  with a positron range in water of 2.9 mm (maximum) and 0.64 mm (mean) that is comparable with that of 18F (≈1 mm), the isotope used in 18F-FDG PET. This is an essential advantage of 64Cu compared with 68Ga. In addition, 64Cu has a half-life of 12.7 hours giving the opportunity for delayed imaging. We found higher 64Cu-DOTATATE uptake in the index vessel compared with the contralateral carotid artery in our population; however, most included patients also had significant plaques in the contralateral vessel. We therefore suggest that it is reasonable to expect an even higher absolute difference in SUVmean values between atherosclerotic versus healthy carotid arteries. Importantly, we found that plaque burden was not correlated with plaque 64Cu-DOTATATE uptake, indicating that plaque macrophage activity is not associated with plaque size per se. These results emphasize that PET has a promising role in molecular characterization of vulnerable plaques by providing in vivo information. We used hybrid PET/MRI instead of PET/CT in our study. MRI has superior soft tissue contrast allowing for better delineation of the carotid artery and atherosclerotic plaque when compared with CT.9 Finally, our PET/MRI system acquires the PET and MRI simultaneously allowing for perfect alignment between the 2 sets of images, when compared with the sequential acquisition in PET/CT where minor head movements can cause misalignment.Only a few studies have used somatostatin receptor imaging in atherosclerosis.13 which is why we use it as a surrogate measure of macrophage load. Several previous studies have shown a good correlation between CD68 and uptake of the glucose analogue FDG.16 Because FDG can be labeled with a positron emitter, this compound can be used for in vivo imaging. However, the major drawback of this imaging tracer is the low specificity of FDG limiting its use in assessing vulnerability of plaques.CD68 is a class D scavenger receptor and, although not exclusively expressed by macrophages, it is a widely used macrophage marker,17 A crucial role of hemorrhage-associated macrophages is to clear hemoglobin–haptoglobin complexes directly via the CD163 receptor and reduce oxidative stress, which subsequently mediates anti-inflammatory properties in vulnerable atherosclerotic plaques.19 Macrophages expressing CD163 have been detected in atherosclerotic plaques,20 and the soluble form of CD163 is associated with coronary atherosclerotic burden in the general population.21 The association between expression of CD163 and in vivo imaging of atherosclerotic plaques has not been investigated previously. However, a study of HIV infected patients found a correlation between plasma concentration of soluble CD163 and FDG uptake in the ascending aorta.22 Interestingly, a recent study into M1/M2 polarization of macrophage phenotypes in carotid plaques found that M2 macrophages are present in both symptomatic and asymptomatic plaques, whereas M1 macrophages are exclusive to symptomatic plaques.23CD163 is a hemoglobin scavenger receptor and macrophage-specific protein. It is upregulated in a subpopulation of alternatively activated M2 macrophages called hemorrhage-associated macrophages that are found in hemorrhagic zones of atherosclerotic plaques.18 Little recent work has been done on TNF-α expression in atherosclerotic lesions in humans; however, one investigation reported markedly raised (but nonsignificant) TNF-α RNA levels in symptomatic human endarterectomy specimens,24 and its role in systemic inflammatory disease is inarguable.26TNF-α is a cytokine involved in systemic inflammation and an important part of the acute phase reaction. It is produced primarily by activated macrophages in inflammation and considered a principal marker of M1 activation.3 were not associated with 64Cu-DOTATATE uptake, highlighting how the 2 tracers image different biological processes using PET. Our finding substantiate that 64Cu-DOTATATE is correlated primarily with CD163 positive macrophages (CD163+) and only weaker with CD68 positive macrophages (CD68+) making 64Cu-DOTATATE uptake a predictor of (hemorrhage-associated macrophages) macrophage activity. Considering our selection of molecular M1/M2 macrophage-polarization markers (TNF-α/CD163), these results indicate that 64Cu-DOTATATE PET detects an M2-subset–driven macrophage response. Although we did find a good correlation between CD68 and CD163 gene expression  of CD68/CD163 expression as the tissue slices were homogenized in toto in the mRNA isolation procedure. PET imaging with 64Cu-labeled DOTATATE is not equivalent to PET imaging with 18F-FDG as previously shown.7 The exact role of different macrophage subsets including CD163+ macrophages of the hemorrhage-associated phenotype in atherosclerosis needs further elucidation, and future studies will have to show whether 64Cu-DOTATATE has higher specificity than 18F-FDG for imaging vulnerability of plaques.Molecular markers of plaque vulnerability  that were previously found to be associated with FDG-uptake18F-FDG where imaging should be postponed to ≈3 hours after injection because FDG uptake in the plaques is either stable or increases, whereas FDG is cleared from the blood leading to improved target-to-background ratios.29 As FDG, in contrast to 64Cu-DOTATATE, is not bound to membrane-bound receptors but is continuously taken up and trapped by the cells, this discrepancy in optimal timing of imaging is not surprising. It could be speculated that instability of the 64Cu–DOTA complex in vivo33 and thereby increase in nonspecific background could be an explanation for the lack of correlation at the late time-point. However, because there was no increase in background activity observed, this explanation seems unlikely. Also, we recently published the first clinical PET study using the tracer 64Cu-DOTATATE in patients with neuroendocrine tumors. Here, we scanned patients 1, 3, and 24 hours post injection and did not see large liver accumulation or increase in blood-borne activity indicating that 64Cu-DOTATATE is indeed stable in humans for a long time.8 Also, as part of the approval by the Danish Health Authorities of 64Cu-DOTATATE, extensive stability studies were undertaken, demonstrating a shelf-life of at least 24 hours. Taken together, we find it most likely that decreasing signal-to-noise ratio over time explained by decrease in signal from receptor binding fully explains why early scans are superior. Based on our results, we therefore suggest that PET imaging of atherosclerosis with 64Cu-DOTATATE should be performed early, for example, at 80 minutes after injection.We found only a moderate correlation between tracer accumulation at the early versus the later scan, and the significant association between tracer uptake and macrophage markers (CD68 and CD163) disappeared at the late time scan. In every single case, the SUV value decreased from early to late imaging indicating that binding to somatostatin receptor subtype-2 decreases over time. This is in contrast to PET imaging of atherosclerosis using 64Cu-DOTATATE seems promising, it is inherently challenged by low tracer uptake and a poor spatial resolution that could limit the usefulness. Nevertheless, the spatial resolution may not be a major obstacle, as it is at the same level as for FDG-PET, which increasingly seems to establish itself as an accepted method for plaque evaluation. However, only large clinical studies, ideally with head-to-head comparison with other methods, can establish the true clinical value of our method and its relation to other methods, for example, FDG-PET.Although our imaging method using 64Cu-DOTATATE, in human atherosclerotic plaques. We found a correlation between in vivo tracer uptake and ex vivo markers of activated macrophages. This association could potentially improve noninvasive identification of vulnerable plaques.In conclusion, we demonstrate the uptake of a novel PET tracer, The expertise and technical support of principal technicians Karin Stahr and Jakup Poulsen with positron emission tomography (PET)/MRI procedures and PET reconstructions are highly valued as is the expert statistical advice by Julie Lyng Forman.The unrestricted financial support from the Danish Heart Foundation, the Research Foundation of Rigshospitalet, the Danish Medical Research Council, and the John and Birthe Meyer Foundation is gratefully acknowledged. The PET/MRI scanner was donated by The John and Birthe Meyer Foundation.None


Different viruses are detected in the cerebrospinal fluid (CSF) of HIV-positive patients, with controversial implications in central nervous system (CNS) impairment. Objectives: to evaluate the presence, frequency and associations of Epstein-Barr virus (EB) in the CSF in HIV-positive patients.We retrospectively analyzed CSF samples from HIV-positive adult patients (≥18 years old), with or without CNS impairment, collected between Oct 2011 and Oct 2012 in a Romanian tertiary infectious diseases hospital.We performed a multiplex PCR coupled with electrospray ionization – time-of-flight mass spectrometry (Abbott Molecular) which can simultaneously detect: herpesviruses (1-5 and 8), polyomaviruses, enteroviruses, adenoviruses and parvoviruses.The patients were characterized based on the immunological and virological HIV status, neurological findings (including neuroimagery and CSF exam) and comorbidities.A number of 55 patients were analyzed, with a mean age of 33.4 years (31.5 median) and a male:female ratio of 1.9:1. The CD4 count had a mean of 32 (75 median and IQR=225).The most frequently detected virus was EB in 20/55 cases.The EB-positive subgroup had a similar mean age of 33.5 years (median of 31.0) but a different male:female ratio, of 1.2:1. The CD4 count had a mean of 105.7 (59.5 median and IQR=156).The following analyses refer to the EB-positive group: 17/20 patients had neurologic impairment. Imagery was performed in 15 cases and was normal in 5. CSF cellularity had a median of 2/cmm, IQR=10, CSF-glucose a median of 48.0 mg/dL, IQR=34 and CSF-protein a median of 55.5mg/dL, IQR=49.5.Mycobacterium tuberculosis (2 cases), Cryptococcus neoformans (2 cases) and Toxoplasma gondii (one case).In 8/20 cases EB was found as singular agent in the CSF and in 12/20 it was associated with other microorganisms: other herpesviruses (3 cases), JC virus (4 cases), The CSF-HIV viral load was available in 12/20 cases, being detectable in 10 cases.Regarding the neurological events , EB was probably causal in 4/20 cases, possibly causal in another 4/20 cases and was a bystander in 12/20 (3 cases with no impairment and 9 cases in which the impairments were strongly attributable to another germ).EB is the most frequent agent found in the CSF (40% of HIV-positive cases), mainly in women. It can be mono-detected but it especially appears in multiple associations. In more than half of the cases EB acts as an innocent bystander.


The strict relationship between shunt-responsiveness and increased resistance to CSF outflow (Rout) as reported in 1981 by Borgensen and Gjerris  and rece310 adult patients (age 40-86) were included in retrospective analysis. All patients had probable NPH following clinical assessment including imaging. Patients underwent lumbar or intraventricular infusion studies and were available for follow-up via the multidisciplinary CSF clinic. Outcomes were assessed using the in-house Cambridge Outcome Scale; a pragmatic categorization of patient cohorts into three groupings – sustained improvement, short-term improvement and no improvement. Compensatory parameters were calculated on the basis of infusion test: baseline ICP and pulse amplitude, Rout, elasticity, estimator of CSF production, slope of amplitude-pressure regression line. Consultants deciding about shunting were not blind to results of infusion study.79% of patients showed improvement . Improvement rate increased from 1992 (60%) to 2013 (86%); p=0.0003. Of all calculated CSF compensatory parameters, only Rout was associated with outcome (p=0.014). Pulse amplitude of ICP (peak-to-peak), slope of amplitude-pressure regression line, elasticity, did not correlate with outcome significantly. Patients with Rout>13 mm Hg/(ml/min) had an improvement rate of 79%, compared to 63% (p=0.011) with Rout<13. Notably, none of patients with low Rout (lower than 6 mm Hg/(ml/min); N=5) improved after shunting. Neither age nor sex correlated with outcome.Rout was the only CSF compensatory parameter correlating with outcome following shunting. The relationship was weak but significant. Infusion studies appeared to be helpful in the assessment of compensatory parameters both for diagnostic and to yield baseline values as a benchmark for further investigations in cases of suspected shunt malfunctions and complications.


Classical midazolam–opioid combination for gastrointestinal endoscopy sedation has been adopted for decades. Dosing regimens have been studied but most require fixed dosing intervals. We intend to use a sophisticated pharmacodynamic tool, response surface model (RSM), to simulate sedation using different regimens. RSM can predict patient's response during different phases of the examination and predict patient's wake-up time with precision and without the need for fixed dosing intervals. We believe it will aid physicians in guiding their dosing strategy and timing.The study is divided into 2 parts. The first part is the full Greco RSMs development for 3 distinct phases: esophagogastroduodenoscopy (EGD), colonoscopy, and intersession (the time lapse between procedures). Observer's Assessment of Alertness Score (OAA/S) is used to assess patient response. The second part simulates 6 regimens with different characteristics using the RSMs: midazolam only, balanced midazolam and opioids, high-dose opioids and midazolam, low-dose midazolam with high-dose opioids, high-dose midazolam and low-dose opioids, and finally midazolam with continuous opioid infusion. Loss of response at 95% probability for adequate anesthesia during examination and return of consciousness at 50% probability during intersession was selected for simulation purposes.2. About 56.7% were females and none received prior abdominal surgery. The cecal intubation rate was 100%. Only 1 patient (3%) developed temporary hypoxemia, which was promptly managed with simple measures. The RSMs for each phase showed significant synergy between midazolam and alfentanil. The balanced midazolam and alfentanil combination provided adequate anesthesia and most rapid return of consciousness. The awakening time from the final drug bolus was 7.4 minutes during EGD and colonoscopy stimulation, and 9.1 minutes during EGD simulation.The average age of the patient population is 49.3 years. Mean BMI is 21.9 ± 2.3 kg/mSimulation of regimens with different characteristics gives insights on dosing strategies. A balanced midazolam–alfentanil regimen is adequate in providing good anesthetic depth and most rapid return of consciousness. We believe with the aid of our RSM, clinicians can perform sedation with more flexibility and precision. Good instrumentation conditions as well as less drug side effects can be achieved by combining an opioid and a hypnotic agent.1 Recent American and European guidelines focused on the use of propofol8 and works regarding the use of propofol have been vigorously done.10 However, propofol has a narrower therapeutic window and may increase the chance of hypotension or respiratory depression that requires intervention as compared with the use of midazolam and alfentanil.11 There are significantly less works emphasizing the dosing strategies using a benzodiazepine and an opioid, probably because they possess more complicated pharmacokinetic properties.12 Alfentanil is a good opioid choice for gastrointestinal endoscopies because of its quick onset time (1–2 min after intravenous injection), short t1/2ke0 resembling thiopental,13 and short duration of effect lasting approximately 10 minutes.13 There are also interesting works done by Short et al14 indicating stronger synergism between midazolam and alfentanil than propofol and alfentanil.16 Lower potency opioids such as meperidine, tramadol, or morphine or adjuvants such as clonidine are generally not used during outpatient procedures. The longer duration of action for these drugs may raise concerns about respiratory function or blood pressure when discharging the patient.Gastrointestinal endoscopy sedation should be characterized by rapid induction of sufficient amount of anesthetics and rapid awakening. Optimal dosing regimen selection in literatures focused on the quality of sedation, occurrence of side effects, and time to awakening among different anesthetic individuals.18 A 2-drug model is usually used because triple or more drug interactions increase model complexity significantly and is less comprehensible graphically. A surface will encompass the complete set of isobolograms, concentration–effect curves, and curve shift effects. The RSMs are most often used clinically to predict wake up time,19 tolerance to stimuli,22 or occurrence of side effects.23 We can help define the optimal dosing regimen by analyzing different but relevant surfaces simultaneously. Another feature that makes RSMs stand out from other models is that it does not require a fixed dosing interval and can cope with very variable examination time.Giving 2 drugs simultaneously requires the knowledge of how they interact with one another. Response surface model (RSM) is a sophisticated pharmacodynamic tool for analyzing drug interaction and responses for multiple drugs.This study is designed to tackle several issues. First, traditional dosing regimen studies require a fixed dosing timetable and examination time. Second, very little work has been done for the benzodiazepine-opioid RSM. Our study is divided into 2 parts. The first part of our study involves building the model for simulation. The second part of the study deals with different midazolam and opioid simulations through RSM, which was developed in the first part, to rationally delineate dosing strategies for gastrointestinal endoscopy.Patients between 18 and 65 years old, American Society of Anesthesiologists (ASA) Physical status I ∼II who underwent EGD (esophagogastroduodenoscopy) and colonoscopy as a single-stage procedure, were chart-reviewed. Those with documented verbal communication impairment, cerebrovascular diseases, incomplete records, or a history of sedative, opioid, or chronic alcohol use were excluded. Requirement for written informed consent was waived by the Institutional Review Board (IRB 2014-12-001BC) of Taipei Veterans General Hospital. After setting up the patient on standard anesthetic care monitoring, continuous 3 L/minute supplemental oxygen was given through nasal cannula to maintain oxygen saturation greater than 90%. One anesthesiologist performed sedation using midazolam and alfentanil. The induction dose of midazolam and alfentanil was 0.03∼0.04 and 6∼9 mg/kg, respectively. A single gastroenterologist performed the examinations.25 We define OAA/S < 2 as loss of response (LOR) in EGD and colonoscopy, while OAA/S < 4 indicates LOR during intersession. The plasma drug concentrations are calculated using TIVAtrainer . Opioid concentrations are converted to alfentanil equivalents using a potency ratio .17 The Greco RSM construct is selected for model building (Equation 1)26 The examinations were divided into 3 distinct phases for model development: EGD, colonoscopy, and intersession (the time lapse between the procedures). Patient responses were evaluated with the Observer Assessment of Alertness/Score (OAA/S) Table .24,25 We50 is the value in which the concentration of the drug is required to exert 50% maximal effect alone. The parameter α is the interaction relationship between midazolam and alfentanil, and γ is the steepness that describes the descent of the surface. Interaction is synergistic when α > 0, infra-additive when α <0, and additive when α equals 0. The RSM is an extension of the Hill equation. The resulting 3-dimensional surface theoretically holds the entire drug interaction relationship for the given endpoint, in this case LOR. Single or multiple isoboles or concentration-effect curves can be extracted from the surface to work with. We used Matlab  software to perform model fit and parameter estimation. Bootstrap-based programming was written with 1000 iterations for model stability. Parameters for each of the 3 phases are obtained for simulation analysis.where E is the probability of LOR and ranges from 0 to 1. Plasma concentration of the drug is expressed as Cp. CpDosing regimens are searched in PUBMED using the keywords: midazolam, alfentanil, fentanyl, remifentanil, colonoscopy, and gastrointestinal endoscopy. Studies with insufficient dosing details, pediatric patients, geriatric patients, or use of premedications were excluded. Regimens with specific characteristics are sought: midazolam only, balanced midazolam and opioids, high-dose opioids with high-dose midazolam, low-dose midazolam with high-dose opioids, high-dose midazolam and low-dose opioids, and finally midazolam with continuous opioid infusion. Pure opioid regimens were not candidates because our endpoint requires the patient to be unresponsive. Two sets of simulation will be performed, EGD alone and EGD immediately followed by colonoscopy. The 50% probability is the targeted value for model-predicted return of consciousness. Adequate anesthesia occurs at our defined 95% probability of LOR. Our main goal is to find an optimal dosing regimen that will provide adequate anesthesia and the most rapid return of consciousness.Forty patients were enrolled, but in the end only 33 were eligible for model building. Of the excluded ones, 5 had an incomplete record and 2 had known cerebrovascular disorders. The patient demographics are summarized in Table The cecal intubation rate was 100%. During the procedures, only 1 patient developed hypoxemia that required intervention. Pulse oximetry displayed saturation between 80% and 90% for approximately 2 minutes. Chin-lift maneuver with increased oxygen flow through nasal cannula increased the patient's saturation above 90%. There were no subsequent events.50alfs were all very large and not achievable with normal dosing during these procedures. This is consistent with the fact that opioids do not produce hypnosis well.The number of concentration pairs available for pooling and model building were 68, 75, and 75 for EGD, colonoscopy, and intersession, respectively. Total midazolam dosage was 0.049 ± 0.016 and 0.013 ± 0.005 mg/kg for alfentanil. The required model parameters (for Equation 1) are summarized in Table 2 as our simulation patient according to our patient demographics. Slight modifications from the original identified regimens had to be made to define specific dosing intervals and dosages for simulation , and a drug washout period of 15 minutes in EGD with colonoscopy and 25 minutes in EGD only. The intersession RSM is applied to the induction and washout phase. The general rule is that midazolam is given at the beginning of the induction because the onset time is approximately 3 minutesThe simulation results for the 5 regimens are illustrated in Figure This simulation is done without colonoscopy and further highlights the need for a very rapid increase in anesthetic depth and return of consciousness. Only regimen 1B and 5 reached adequate anesthesia Figure  during EOur study has graphically illustrated how regimens with different characteristics perform during gastrointestinal procedures. Midazolam and opioids together are capable of providing reasonably rapid return of consciousness, and can be predicted by RSMs. Regimen 1B with balanced midazolam and alfentanil administration consistently resulted in adequate anesthesia and more rapid recovery.33 Concurrent administration of a hypnotic agent and an opioid gives better instrumentation condition 4 safety and higher patient satisfaction.35 Bannert et al36 reviewed 52,506 colonoscopies and concluded that sedation increased cecal intubation rate in both men and women, but the effect is more pronounced in women. Midazolam–opioid combination reduced the rate of hypoxemia as compared with midazolam alone. A meta-analysis reported hypoxemia in 18% of the patients when midazolam is used alone, and 6% when midazolam is combined with an opioid.10 In our patients, only 1 (3%) developed hypoxemia requiring intervention. When compared with propofol, midazolam may result in less hypoxemia events and require less anesthesiologist interventions.37The combination of midazolam and an opioid for procedural sedation is widely used and provide faster return of consciousness than diazepam.38The cecal intubation rate was 100% in our patients and there is a low chance of respiratory depression (3%) during anesthesia. Model built from this population implies that the RSM can facilitate colonoscopies while avoiding unwanted respiratory depression. Hemodynamic stress from endoscopy pain can contribute to myocardial ischemia. Sedation offers additional benefits and a prospective study concluded that sedation with midazolam and propofol may provide protection against stress response.35 During colonoscopy, higher pushing force is at the sigmoid colon and this part of the colon is associated with the highest perforation rate (0.045%).39 Another study used magnetic endoscope imaging also reported pain most commonly occurred at sigmoid colon (77%), while the proximal colon was infrequent (4%).40 It would seem unnecessary to keep the same anesthetic depth in the second half of colonoscopy as the beginning.In Figure 10 Most hypnotic agents such as propofol and midazolam lack analgesic property. It would require very large doses of hypnotics to cover the pain encountered during endoscopies, along with a higher chance of unwanted side effects and poorer instrumentation conditions.41 Inadequate anesthetic depth in regimen 2 and regimen 4 implies that neither low opioid–high midazolam or low midazolam–high opioid combinations are regimens of choice.Regimen 1A produced very poor LOR and reflects the fact that sedation with a single agent should be avoided. Many sedation regimens use single agents for comparison, either at low or high doses.Regimens 3 and 5 tend to overshoot drug concentration required to produce adequate anesthesia Figure . It takeRegimen 1B has also been identified as the better performing technique, with adequate anesthesia and faster return of consciousness. The total midazolam used was 4 mg and alfentanil was 600 μg. With this dosing, anesthesia is anticipated to last for 9.1 minutes  performs better in terms of providing adequate anesthesia as well as the most rapid return of consciousness. Although our model can be used by any field of expertise, it is generally accepted that sedation specialists such as anesthesiologists who are airway experts be present while performing sedations. We believe with the aid of our RSM, clinicians can perform sedation with more flexibility and precision.