Trachoma is the leading infectious cause of blindness worldwide. Corneal scarring, which causes trachoma-related blindness, occurs when the upper eyelashes are turned inward and rub on the eye (cornea). This is called trichiasis, and if the lid margin turns inward, the term entropion is used. Currently, there are an estimated 8.2 million people with trichiasis and 3.1 million people are blind from trachoma.A systematic review of population-based trachoma surveys has shown that women are affected by trichiasis approximately twice as often as men. Trichiasis is more common with increasing age; however, in communities with very high levels of trachoma infection, trichiasis can occasionally occur in children.Persons who develop trachomatous trichiasis (TT) usually need treatment to either surgically turn the eyelashes outward from the eye or to remove one or two in-turning eyelashes which are not central or touching the cornea; the latter is pulling out the in-turned lash or lashes with forceps, a procedure called epilation. Bilamellar tarsal rotation (BLTR) or posterior lamellar tarsal rotation (PLTR) are procedures widely used in trachoma endemic countries to surgically treat TT and are believed to produce comparable results.In some countries, there are huge numbers of persons with untreated TT, often living in poor and remote communities. For example, in Ethiopia, there are an estimated 1.2 million people with TT who need an operation. However, the number of TT operations currently performed each year in Ethiopia is about 80,000. At this rate, it will take 15 years just to clear the backlog, without considering any new cases which will occur!There are several reasons for the large numbers of untreated trichiasis patients in endemic countries:Patients may be unaware that surgery can help, or they may be afraid of an eye operation; as a result, uptake of TT surgery is often low, even when surgery is provided free of charge. Sometimes, the fear is reinforced by awareness in the community that, at a particular clinic, trichiasis often comes back after surgery, which has a negative impact on uptake.Some patients find the cost of travel to seek eye services, or the lack of a companion to go with them, to be a significant obstacle. This is particularly true for women who also have to look after children and the household and cannot afford the time to go for treatment. Sometimes, it is just too great a distance to a health facility, so people will not go for treatment.In some situations, services are simply not available, nobody has been trained to perform trichiasis surgery, or the necessary equipment and consumables are not available.Strategies to address the TT backlog will vary from country to country and setting to setting.These may include: creating awareness of treatment for trichiasis through health education, including radio programmes; ensuring TT surgery is available at low cost and close to where people with TT live; or conducting enhanced outreach in communities where trachoma is common.TT surgery can be performed by well-trained ophthalmic nurses, assistants, or doctors. There is good evidence that TT surgery can be done by non-ophthalmologists with comparable results to those of ophthalmologists.One of the challenges is to encourage the trained TT surgeon to continue to work in rural areas and to equip them so that they are able to perform sufficient TT operations per year to maintain good experience and quality.Unfortunately, the quality of training of TT surgeons can be variable and adequate supervision may be lacking, leading to high rates of recurrence of TT after surgery. Surgeons who only do a small number of TT operations each month tend also to have poor surgical outcomes, leading to a vicious cycle of low uptake, low productivity, and poor surgical quality and outcome.Breaking this cycle requires good planning and a willingness to acknowledge that results can be improved.In order to develop a TT service it is useful to address various levels of eye care delivery.At a national level, it is necessary to identify areas with a high prevalence of TT and to prioritise these areas for TT surgery programmes.TT surgeons need to be given good quality training and be adequately equipped. Quality of care is essential. In order to improve the quality of surgery, training of TT surgeons should be standardised and surgeons should be certified using the World Health Organization manual on assessment of trichiasis surgeons.Due emphasis should be given to the selection of trainees, the creation of a career pathway, and supervision of TT surgeons.TT surgeons must also have adequate supplies of instruments and consumables.Services, whether static or outreach, that are staffed with poorly skilled, inadequately supervised TT surgeons can result in poor surgical outcomes and negative publicity for the programme.At the health centre level, transport to provide outreach programmes for TT surgery in affected communities is required, together with good provision of consumables to perform the operation: medicines, sutures, dressings, and so on.It may be necessary, in some situations, to consider offering incentives  to encourage good TT surgeons to work in high-volume TT programmes in remote areas.At the community level, women must be specifically and deliberately targeted for trichiasis surgery. A successful trachoma programme requires the involvement of affected communities through their village leaders, women's group leaders, teachers, community health agents, health extension workers, or similar frontline health personnel. Recruiting village women who have had successful TT surgery to raise awareness and encourage others with TT to have the operation has proven to be a successful strategy.An outreach programme includes awareness creation in the community about trachoma, seeks community involvement in planning and executing the activities, and tries ultimately to engage the community to such an extent that they become the true owners of the programme. Community involvement and engagement is therefore essential for community ownership and the successful implementation of the full SAFE strategy.Community and local health service planners need to decide what contribution community members can make towards the cost of surgery. This will help to achieve sustainable service delivery and avoid a situation where the community either undervalues the service (perhaps because they consider a free service to be inferior) or where a state of dependence is created. Having said that, the cost of surgery should not be so high that poor patients cannot afford it.In summary, it is essential to consider patients' needs. This will require comprehensive planning at national, district, and community levels to adapt and strengthen the health system to meet these needs. The ultimate aim is for patients with TT to have successful surgery, be satisfied with the result, and be advocates in their communities. Only then will we achieve the ultimate goal of the elimination of blinding trachoma.


To determine the relationship between biochemical markers of myocardial injury in acute myocarditis and the cardiac MR imaging features.Cardiac MR criteria for acute myocarditis (“Lake Louise Criteria”) include scarring as defined by high-signal-intensity areas in late gadolinium enhancement (LGE), and inflammatory markers as defined by an increased early contrast uptake  and edema . Troponin is a widely used clinical marker for cardiomyocyte death; however, the relationship between biochemical markers of myocardial injury and these imaging features has not been clearly established.Thirty-one patients who had troponin-T levels measured at presentation and had the diagnosis of acute myocarditis based on clinical factors and CMR criteria “Lake Louise Criteria” were evaluated. MR images were assessed for the volume of LGE, %EGE and EGE ratio, as well as T2 signal intensity ratio normalized to skeletal muscle. Ordinary least squared linear regression was used to determine the relationship between these imaging features and peak serum troponin-T concentration in the acute presentation.2= 0.57, Beta coefficient= 16.8, p<0.001) and , respectively. The T2 signal intensity ratio and EGE ratio did not show a relationship to troponin levels  and , respectively.There was a linear increase in the total volume of LGE and EGE with increases in square root troponin-T concentration (RIn the setting of acute myocarditis troponin-T concentrations show the strongest correlation with LGE and, to a lesser degree with EGE values. There is no correlation to the calculated EGE ratio or T2 ratio. These finding suggest that LGE specifically reflects irreversible myocardial injury, whereas other CMR criteria appear to reflect processes that are not associated with myocardial necrosis.


Background. Although CSF cytology and MRI are standard methods to diagnose neoplastic meningitis (NM), this complication of neoplastic disease remains difficult to detect. We therefore reevaluated the sensitivity of gadolinium (GD)-enhanced MRI and cerebrospinal-fluid (CSF)-cytology and the relevance of tumor type and CSF cell count. Methods. We retrospectively identified 111 cases of NM diagnosed in our CSF laboratory since 1990 with complete documentation of both MRI and CSF cytology. 37 had haematological and 74 solid neoplasms. CSF cell counts were increased in 74 and normal in 37 patients. Results. In hematological neoplasms, MRI was positive in 49% and CSF cytology in 97%. In solid tumors, the sensitivity of MRI was 80% and of cytology 78%. With normal CSF cell counts, MRI was positive in 59%  and CSF cytology in 76% . In cases of elevated cell counts, the sensitivity of MRI was 72%  and of CSF cytology 91% . 91% of cytologically positive cases were diagnosed at first and another 7% at second lumbar puncture. Routine protein analyses had a low sensitivity in detecting NM. Conclusions. The high overall sensitivity of MRI was only confirmed for NM from solid tumors and for elevated CSF cell counts. With normal cell counts and haematological neoplasms, CSF-cytology was superior to MRI. None of the analysed routine CSF proteins had an acceptable sensitivity and specificity in detecting leptomeningeal disease. Neoplastic meningitis (NM) is a diffuse spreading of malignant cells into the cerebrospinal fluid (CSF) and/or into the adjacent leptomeninges. The CSF flow can distribute the malignant cells throughout the entire central nervous system (CNS) causing a variety of clinical symptoms. Early diagnosis of NM is regarded to be crucial since the rapid institution of therapy may offer the best chances for successful treatment –3.In autopsy series of patients with malignant disease, leptomeningeal involvement was diagnosed in up to 19% . Even inIn solid tumors, NM occurs in about 5–8% of patients, most commonly with carcinomas of the breast and lung and melanoma –3. AmongNeoplastic meningitis may occur diffusely with malignant cells floating freely within the CSF or as adherent types . While CThe archives of the Laboratory of Neurochemistry, the Department of Neuroradiology, and the general diagnostic database of the University Medicine Göttingen were searched for patients with neoplastic meningitis of which both CSF cytology and MRI of the CNS were available. Only cases with complete documentation of the radiological and cytological examinations were included. Between 1994 and 2009, 111 evaluable cases of NM were identified.The diagnosis of NM was proved by review of the cytospin slides  and MR-images  and review of the clinical files with regard to clinical symptoms and further clinical course. T1-weighted images with and without gadolinium and FLAIR-sequences were used for the reevaluation of the MR-imaging. Radiological criteria of NM were signal alterations of the meninges and contrast enhancement lining the CSF spaces. Only irregular or nodular contrast enhancement of the meninges was considered suspicious of NM in order to avoid misdiagnosing fine linear meningeal enhancement which is known to occur as an unspecific finding after lumbar puncture –12. CytoFor each case, the diagnostic method leading to the correct diagnosis  was assessed. In addition, the number of CSF probes necessary to establish the cytological diagnosis was analysed. The diagnostic sensitivity was calculated as percentage of positive diagnoses of the total number of cases. This was calculated separately for each diagnostic method in the whole series and for subgroups with normal and elevated cell count and with haematological or solid neoplasms.Between 1994 and 2009, 111 cases of NM were identified: 37 patients had hematological and 74 solid neoplasms. CSF cell counts were elevated in 74 and normal in 37 patients.In the whole series, cytology was significantly more sensitive than MRI . In partCSF cell counts influenced the overall diagnostic sensitivity considerably: CSF-cytology detected neoplastic meningitis more often in cases with CSF pleocytosis than with normal cell counts. Equally, MRI was more often positive with elevated than with normal cell counts. In haematological neoplasms with higher cell counts, CSF cytology remained to be significantly more sensitive than MRI. Of note, we could not detect MRI-positive, cytology negative haematological NM with elevated cell counts in our databases. Thus, we consider the sensitivity of cytology in this specific subgroup to be artificially high.In the whole series with positive CSF cytology, 91% were already detected with the first lumbar puncture. The second CSF sample revealed additional 7% of cases and the third and all additional punctures added only 2% more positive cases. In 11 patients, the first CSF probe was cytologically negative, and no further CSF samples were taken because MRI and clinical symptoms clearly indicated NM. This translates into an overall sensitivity of 79% of all cases with the first puncture, 85% with the second and 86% with three or more samples.Alterations of CSF proteins were variable : total pAlthough a well-known complication of neoplastic disease, the early diagnosis of neoplastic meningitis is often difficult even in these times of modern diagnostic methods. This retrospective analysis found comparable sensitivities of CSF cytology and MRI for solid neoplasms, but a superiority of CSF cytology in hematological malignancies. CSF pleocytosis largely influenced the diagnostic accuracy. The analysis of standard CSF proteins has no positive diagnostic value, but negativity of all standard parameters  is associated with a probability of over 80% that NM is excluded.solid tumors has increased in recent years [While the occurrence of CNS relapse in hematological malignancies has dropped considerably after the introduction of CNS prophylaxis, , 15 the nt years . This isnt years . As a rent years .Since early diagnosis and onset of treatment are regarded to be crucial for successful treatment and prevention of permanent neurological sequelae,  the earlThe analysis presented here proves that in fact MRI is highly sensitive in detecting NM from solid neoplasms, especially in cases of elevated cell counts. In line with other studies, , 24 we oIn hematological malignancies, CSF-cytology was clearly more sensitive in detecting NM than MRI. The rate of 100% sensitivity in cases of elevated cell counts most probably is overestimated because no cytology-negative cases could be identified in this subgroup in spite of extensive search of the databases. In any case, the difference to MRI sensitivity is striking and in line with the findings of other groups. In a large series of primary CNS lymphoma, Fischer et al. found an even lower rate of 4–18% of cases identified by MRI , 26. SimIn addition to the results of previous series, we demonstrate here that the diagnostic sensitivities are considerably higher when the CSF cell count is elevated . Since oAnother scope of this study was to analyse how many CSF samples are needed to achieve an accuracy of CSF cytology of more than 90% as reported previously  Figure . In thisAs expected, the overall cell count and biochemical CSF analysis were unspecific and not suitable to detect NM. Surprisingly, one-third of the cases of NM reported here had normal CSF cell counts. This demonstrates that normal CSF cell counts cannot exclude malignant CSF disease. Interestingly, however, only 16% of all cases had completely normal standard CSF parameters, including total cell count, total protein, lactate, ferritin, and oligoclonal IgG.We conclude that MRI is highly sensitive in detecting NM only in solid, but not in hematological malignancies. Since NM can occur with either cranial or spinal preference, high-resolution imaging of the whole crane and spine is mandatory to localize the spatial distribution of the disease. In addition, MRI is necessary for the detection of solid CNS metastases. CSF cytology can enhance the diagnostic accuracy by 12 to 50% depending on the diagnostic subgroup. In addition, repeated CSF analyses are suitable for the monitoring of treatment effects. Therefore, both methods are needed for the complete staging when NM is suspected.


To describe a case of HTLV-1 associated myelopathy (HAM/TSP) of rapid onset and progression in a renal transplant recipient, and the initial response to empirical treatment.A 50-year-old man was admitted with 12 weeks of evolving spastic paraparesis and urinary–fecal retention, five months after a successful living-donor renal transplantation due to end-stage chronic kidney disease. He was on basiliximab, metilprednisolone, tacrolimus, mychophenolate and prednisone for immunosupression.The physical examination showed pyramidal syndrome predominantly in lower limbs, with hyperalgesia and hypersthesia. The Kurtzke's Expanded Disability Scale (EDSS) showed a score of 7.5. Magnetic resonance imaging showed enlargement and inflammatory pattern in whole spinal cord. HTLV-1 ELISA was positive both in fresh blood and cerebrospinal fluid (CSF). HTLV-1 proviral load by Sybr-Green quantitative real-time PCR performed twice were 1879 copies/10,000 peripheral blood lymphocytes, and 6465 copies/10,000 CSF lymphocytes. Although HTLV-1 screening had not been performed before transplantation neither in the donor nor in the recipient, results were positive in a fresh blood sample from the donor, and negative in one from the recipient cryopreserved two years before transplantation.The patient received empirical treatment with a pulse of corticosteroids, zidovudine, lamivudine, prednisone and baclofen. After one month of treatment, the EDSS showed a score of 7.This case report shows a very acute clinical presentation of HAM/TSP with extensive and diffuse spinal cord involvement in an immunosuppressed patient. In endemic areas, HTLV-1 screening should be included in protocols of organs’ donors and recipients before transplantation.


Chloroflexus aurantiacus is a thermophilic filamentous anoxygenic phototrophic (FAP) bacterium, and can grow phototrophically under anaerobic conditions or chemotrophically under aerobic and dark conditions. According to 16S rRNA analysis, Chloroflexi species are the earliest branching bacteria capable of photosynthesis, and Cfl. aurantiacus has been long regarded as a key organism to resolve the obscurity of the origin and early evolution of photosynthesis. Cfl. aurantiacus contains a chimeric photosystem that comprises some characters of green sulfur bacteria and purple photosynthetic bacteria, and also has some unique electron transport proteins compared to other photosynthetic bacteria.Cfl. aurantiacus has been determined, analyzed and compared to the genomes of other photosynthetic bacteria.The complete genomic sequence of Cfl. aurantiacus to facilitate life under both anaerobic and aerobic conditions, including duplicate genes and gene clusters for the alternative complex III (ACIII), auracyanin and NADH:quinone oxidoreductase; and several aerobic/anaerobic enzyme pairs in central carbon metabolism and tetrapyrroles and nucleic acids biosynthesis. Overall, genomic information is consistent with a high tolerance for oxygen that has been reported in the growth of Cfl. aurantiacus. Genes for the chimeric photosystem, photosynthetic electron transport chain, the 3-hydroxypropionate autotrophic carbon fixation cycle, CO2-anaplerotic pathways, glyoxylate cycle, and sulfur reduction pathway are present. The central carbon metabolism and sulfur assimilation pathways in Cfl. aurantiacus are discussed. Some features of the Cfl. aurantiacus genome are compared with those of the Roseiflexus castenholzii genome. Roseiflexus castenholzii is a recently characterized FAP bacterium and phylogenetically closely related to Cfl. aurantiacus. According to previous reports and the genomic information, perspectives of Cfl. aurantiacus in the evolution of photosynthesis are also discussed.Abundant genomic evidence suggests that there have been numerous gene adaptations/replacements in Cfl. aurantiacus has many interesting and certain unique features in its metabolic pathways. The complete genome may also shed light on possible evolutionary connections of photosynthesis.The genomic analyses presented in this report, along with previous physiological, ecological and biochemical studies, indicate that the anoxygenic phototroph  Chloroflexus aurantiacus was the first filamentous anoxygenic phototrophic (FAP) bacterium  to be discovered  cluster-dependent . It has been suggested that biosynthesis of dNTP is catalyzed by NrdB, NrdJ, and NrdG in aerobic, aerobic and anaerobic, and strictly anaerobic environments, respectively. The activity of NrdJ in Cfl. aurantiacus has been reported [Cfl. aurantiacus genome, suggesting that NrdG and NrdJ produce dNTP for Cfl. aurantiacus in response to the oxygen level  and α-ketoglutarate dehydrogenase , as well as pyruvate:ferredoxin oxidoreductase (or pyruvate synthase)  and α-ketoglutarate:ferredoxin oxidoreductase (or α-ketoglutarate synthase)  are present in the genome (Table coxG and coxSML) have been found in the genome, suggesting that Cfl. aurantiacus can use CO as an electron source during aerobic or semi-aerobic growth. A similar mechanism has been suggested in the aerobic anoxygenic phototrophic proteobacterium Rsb. denitrificans [2 generated from CO oxidation can be assimilated by Cfl. aurantiacus via the autotrophic carbon fixation cycle and/or the CO2-anaplerotic pathways. Under autotrophic growth conditions Cfl. aurantiacus is known to use a unique carbon fixation pathway: the 3-hydroxypropionate (3HOP) autotrophic cycle [2-fixing enzymes , are common to the two pathways.Genes encoding carbon monoxide dehydrogenase , accA , accD  and accB (Caur_3739) [mcr, Caur_2614) [pcs, Caur_0613) [pccB, Caur_2034, Caur_3435), (5) methylmalonyl-CoA epimerase , (6) L-methylmalonyl-CoA mutase (MCM) , (7) succinyl-CoA:(S)-malyl-CoA transferase (smtA (Caur_0179), smtB (Caur_0178)), (8) succinate dehydrogenase , (9) fumarate hydratase , (10) (S)-malyl-CoA/β-methylmalyl-CoA/(S)-citramalyl-CoA (MMC) lyase , (11) β-methylmalyl-CoA dehydratase , (12) mesaconyl-C1-CoA-C4-CoA transferase  and (13) mesaconyl-C4-CoA hydratase   , ATP citrate lyase (the reductive TCA cycle) and acetyl-CoA synthase (the Wood-Ljungdahl pathway) are present, strongly suggesting that these autotrophic carbon fixation pathways are not present in Cfl. aurantiacus.When the genome was first available, some genes required for the 3HOP cycle could not be found , whereasur_3739) , (2) malur_2614) , (3) prour_0613) , (4) protme), phosphoenolpyruvate (PEP) carboxykinase (pckA) and PEP carboxylase (ppc) have been identified, suggesting that Cfl. aurantiacus can assimilate some CO2 and replenish the metabolites in the TCA cycle through the CO2-anaplerotic pathways. The active CO2-anaplerotic pathways have been identified experimentally in other anoxygenic phototrophs during autotrophic, mixotrophic and heterotrophic growth [Cfl. aurantiacus . Moreover, all of the genes in the TCA cycle are present in the Cfl. aurantiacus genome.Additionally, genes encoding malic enzyme  and consumed by malate synthase (EC 2.3.3.9)  in the glyoxylate cycle. With acetate as the sole organic carbon source to support the photoheterotrophic growth of Cfl. aurantiacus, higher activities of isocitrate lyase and malate synthase have been reported [glcDEF, EC 1.1.3.15) and glyoxylate reductase , the glyoxylate shunt and the 3HOP cycle can be employed by Cfl. aurantiacus for assimilating glycolate. Together, genes encoding central carbon metabolism, 3HOP cycle, glycolate assimilation, the glyoxylate shunt and CO oxidation are listed in Table In central carbon metabolism, all of the genes in the TCA cycle as well as the glyoxylate cycle have been identified. The glyoxylate cycle is one of the anaplerotic pathways for assimilating acetyl-CoA, thus lipids can be converted to carbohydrates. Glyoxylate is synthesized and also assimilated in the 3HOP cycle Figure , and is Cfl. aurantiacus does not have genes in the ED pathway, but has genes in the oxidative PP pathway, in agreement with the activities reported for the essential enzymes (glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase) in the oxidative PP pathway [Chloroflexi species . If Cfl. aurantiacus were unable to synthesize FBA, an active pentose phosphate pathway would be required for the interconversion of D-glucose-6-phosphate and GAP, so glucose and other sugars can be converted to pyruvate and other energy-rich species, and vice versa. However, Cfl. aurantiacus has been reported to grow well in glucose and a number of sugars during aerobic respiration [Roseiflexi species , which are closely related to Chloroflexi species, have a putative bifunctional FBP aldolase/phosphatase gene identified [Cfl. aurantiacus genome. Taken together, Cfl. aurantiacus and other Chloroflexi species may employ either a novel FBP aldolase or more than one carbohydrate metabolic pathway. Further efforts will be needed to clarify this picture.Three carbohydrate metabolism pathways are utilized by various bacteria: the Embden-Meyerhof-Parnas (EMP) pathway (glycolysis), the Entner-Doudoroff (ED) pathway, and the pentose phosphate (PP) pathway.  pathway . Genes ipiration , and usepiration . Also, hpiration ,92,93. NCfl. aurantiacus is known to use ammonia as the sole nitrogen source, and several amino acids (nitrogenous compounds), but not nitrate, can enhance the growth. Neither nitrogenase nor nitrogen fixation has been reported in Cfl. aurantiacus [hal), tyrosine phenol-lyase (tpl), asparaginase (aspg), glutamate dehydrogenase (glud1), and glutamate ammonia-lyase (glul), but not nitrogen metabolism (nifDHK) and nitrate reduction, are present in the genome. Note that Cfl. aurantiacus has genes encoding a copper-containing nitrite reductase (EC 1.7.2.1) (Caur_1570) and the α-subunit , but not other subunits (narHIJ) and the catalytic subunit (nasA), of nitrate reductase. Two threonine/serine dehydratases (EC 4.3.1.19), one of which is inhibited by isoleucine may be related to the isoleucine biosynthesis, and other key enzymes in isoleucine biosynthesis have been reported [ilvA genes (Caur_2585 and Caur_3892) encoding threonine dehydratases, and genes in the isoleucine/leucine/valine biosynthesis pathway have been identified . While many organisms reduce APS instead of PAPS to sulfite, it is unknown if Cfl. aurantiacus carries out this reaction as genes encoding APS reductase (EC 1.8.99.2) have not been identified in the genome. In addition to the proposed pathway, sulfotransferase (Caur_2114) can also transfer the sulfate group from PAPS, which serves as a sulfur donor, to an alcohol or amine acceptor for generating various cellular sulfate compounds. PAP is generated as a by-product in the reactions catalyzed by PAPS reductase and sulfotransferase, and has no known functions in metabolism and is likely hydrolyzed to AMP and Pi via PAP phosphatase (unidentified yet). Sulfite reductase  reduces sulfite to sulfide, which is incorporated into cysteine by cysteine synthase A  or cysteine synthase B . The overall proposed sulfur reduction and assimilation pathways are illustrated in Figure c growth ,100. Whereported . SulfateCfl. aurantiacus grows photoautotrophically in the presence of sufficient sulfide [sqr, Caur_3894), has been found in the genome. SQRs belong to the members of the disulfide oxidoreductase flavoprotein superfamily. Other than type II SQRs, type I and type III SQRs with distinct sequences and structures and cofactor requirements have also been reported [2S → EH2(red) + S°), different types of SQR have been identified in various classes of photosynthetic bacteria [Rhodobacter capsulatus ) [Cfl. aurantiacus and cyanobacteria (such as Synechocystis PCC 6803) [Other than using sulfide as the sulfur source during photoheterophic growth,  sulfide -102. Undreported . Althougbacteria : type I,ulatus ) ; type IICC 6803) ; and typCC 6803) . In addiCC 6803) .Cfl. aurantiacus genome. As suggested by phylogenetic analyses [Cfl. aurantiacus and other photosynthetic bacteria. Some proposed lateral gene transfers are listed below and also illustrated in Figure Our paper reports numerous aerobic/anaerobic gene pairs and oxygenic/anoxygenic metabolic pathways in the analyses -8 and coCfl. aurantiacus and the green sulfur bacteria (GSBs). The GSBs have larger chlorosomes and more genes encoding chlorosome proteins [Chlorosomes were transferred between proteins ,29. The acsF gene was transferred either to or from purple bacteria and cyanobacteria, and the bchE gene either to or from heliobacteria, purple bacteria and GSBs.AcsF and BchE are proposed to be responsible for biosynthesis of the isocyclic ring of (bacterio)chlorophylls under aerobic and anaerobic growth conditions, respectively ,32,62-64bc1 or b6/f complex.Two gene clusters of the complex I genes were transferred to (some) purple bacteria. Genes encoding auracyanin may have been transferred either to or from cyanobacteria where the type I blue copper protein plastocyanin is found. Alternative complex III (ACIII) may have evolved from or to the cytochrome 2 fixation, and heliobacteria obtained the gene encoding (Re)-citrate synthase for synthesizing citrate and operating the partial oxidative TCA (OTCA) cycle [Cfl. aurantiacus operates the OTCA cycle, the RTCA cycle in GSBs may have evolved from the OTCA cycle [Genes encoding pyruvate dehydrogenase and α-keto-glutarate dehydrogenase were transferred either to or from purple bacteria and cyanobacteria, and genes encoding PFOR (or pyruvate synthase) and KFOR (or α-ketoglutarate synthase) to or from heliobacteria and GSBs. GSBs may have acquired the ATP citrate lyase gene to complete the reductive TCA (RTCA) cycle for COA) cycle . Since CCA cycle .Chloroflexus aurantiacus is an amazing organism. It has a chimerical photosystem that comprises characteristic types of green sulfur bacteria and purple photosynthetic bacteria. It is metabolically versatile, and can grow photoautotrophically and photoheterotrophically under anaerobic growth conditions, and chemotrophically under aerobic growth conditions. Consistent with these physiological and ecological studies, the Cfl. aurantiacus genome has duplicated genes and aerobic/anaerobic enzyme pairs in (photosynthetic) electron transport chain, central carbon metabolism, and biosynthesis of tetrapyrroles and nucleic acids. In particular, duplicate genes and gene clusters for two unique proteins and protein complexes in Cfl. aurantiacus and several FAP bacteria, the alternative complex III (ACIII) and type I blue copper protein auracyanin, have been identified in Cfl. aurantiacus genome. The genomic information and previous biochemical studies also suggest that Cfl. aurantiacus operates diverse carbon assimilation pathways. In contrast to the purple photosynthetic bacteria, the photosynthetic genes are rather spread out in the Cfl. aurantiacus chromosome. Overall, the genomic analyses presented in this report, along with previous physiological, ecological and biochemical studies, indicate that Cfl. aurantiacus has many interesting and certain unique features in its metabolic pathways.The filamentous anoxygenic phototrophic (FAP) bacteria (or the green non-sulfur bacteria) have been suggested to be a critical group in the evolution of photosynthesis. As the first characterized FAP bacterium, the thermophilic bacterium Chloroflexus aurantiacus J-10-fl was sequenced at the Joint Genome Institute (JGI) using a combination of 8-kb and 14-kb DNA libraries. All general aspects of library construction and sequencing performed at the JGI can be found at http://www.jgi.doe.gov/. Draft assemblies were based on 58246 total reads. Both libraries provided 10x coverage of the genome. The Phred/Phrap/Consed software package (http://www.phrap.com) was used for sequence assembly and quality assessment [Cfl. aurantiacus genome sequence contains 61248 reads, achieving an average of 11-fold sequence coverage per base with an error rate less than 1 in 100,000.The genome of sessment -109. Aftsessment  or transChloroflexus aurantiacus J-10-fl has been annotated by the default JGI annotation pipeline. Genes were identified using two gene modeling programs, Glimmer [The genome of  Glimmer  and Crit Glimmer  as part  Glimmer  with def Glimmer . Genes w Glimmer , and err Glimmer  and GenB Glimmer  and TIGR Glimmer  database Glimmer  using RP Glimmer , TMHMM [ Glimmer , and sig Glimmer . The ann Glimmer .The 16S rRNA gene sequences of various photosynthetic bacteria were obtained from NCBI. The sequences of 16S rRNA genes were aligned using the program Bioedit , and thehttp://www.bacterio.cict.fr/index.html).Three-letter abbreviation for the generic name of phototrophic bacteria follows the information listed on LPSN (List of Prokaryotic names with Standing in Nomenclature), an on-line database curated by professor Jean P. Euzéby (The authors declare that they have no competing interests.BKP and REB designed research and provided DNA; KB, OC, ED, CSH, LJH, MLL, AL, FWL, NM and SP conducted sequencing, assembly and automated annotation; KHT, BMH, LEK and REB analyzed data; and KHT and REB wrote the paper. All authors read and approved the final manuscript.Table S1. Annotation of photosynthetic genes in Cfl. aurantiacus and Cba. tepidum.Click here for file


Renal cell carcinoma (RCC) is considered a contraindication for transplant. However, an increasing number of cases of transplant kidneys with RCC have been reported with encouraging results. We present our experience of two cases of transplanting kidneys with small RCCs. Donors and recipients were aware of the presence and possible consequences of RCC in the transplanted kidney before transplantation. Cases were discussed in the multidisciplinary team meetings. Regular, 6–12 monthly follow-up of donors and recipients was carried out with ultrasonography and/or computed tomography to detect recurrence of RCC or new tumours in the recipients’ transplant kidneys or the donors’ native kidneys. The outcome was recorded. There were no suspicious masses in the any of the kidneys during the follow-up period. The transplant kidneys are functioning. Due to the shortage of organ supply, dependence on live kidney donation is reducing and restrictions for accepting donors are becoming less stringent. Donor safety is nevertheless of paramount importance. Donors therefore undergo extensive evaluation prior to surgery. This includes computed tomography (CT) renography or magnetic resonance angiography (MRA). These investigations lead to increased diagnosis of unexpected pathologies in donors.One of these diagnoses is the presence of small (<4cm) renal cell masses and renal cell carcinomas (RCCs). This is an overview of the management of RCC discovered either during the evaluation process of the kidney donors or after donor nephrectomy in one of the UK major centres of kidney transplantation.A retrospective review of the last five years of living donor kidney transplants was carried out. Two cases with RCC in the transplanted kidneys were identified.In 2006 an ABO incompatible transplant took place between a 45-year-old healthy female kidney donor and her 57-year-old husband with end stage renal disease (ESRD) secondary to Alport syndrome. The donor also had a history of hypertension and post-immunosuppression lymphoproliferative disorder. The recipient had desensitisation with rituximab and plasma exchange according to the local ABO incompatible transplant protocol. During the donor nephrectomy, a 0.5cm cystic lesion was discovered in the lower pole of the kidney. The lesion was excised with part of the surrounding healthy looking tissue and a frozen section procedure was performed, which proved the lesion to be clear cell RCC with a cancer free surgical margin.The case was discussed with the recipient, and the risk of recurrence of the RCC in the transplant kidney and consequences of this were explained thoroughly. The recipient decided to go ahead with the transplant, accepting the risks explained to him.In 2008 a 72-year-old healthy female potential donor was discovered to have a 14mm suspicious exophytic mass at the posterior medial aspect of the upper pole of the left kidney during the donor workup with pre-operative magnetic resonance angiography. CT of the chest, abdomen and pelvis confirmed the presence of the lesion and excluded the presence of any other suspicious lesions.The potential recipient was the donor’s 71-year-old sister with ESRD secondary to focal segmental glomerulosclerosis with a history of a right mastectomy for a breast carcinoma. This was performed five years prior to the transplant and the patient had been free of breast cancer during that period. However, the patient had a B cell positive cross-match, and desensitisation with rituximab and plasma exchange was necessary before the transplant, according to the local protocol.The case was discussed with the donor and the recipient, explaining the possible diagnosis of RCC in the suspicious mass as well as the possible recurrence of the RCC in the transplant kidney and its consequences. Both the donor and recipient accepted the risk and the transplant was carried out.In these two cases, regular follow-up by ultrasonography every 3 months and CT every 6–12 months was carried out for both the donor and recipient for recurrence of the original tumour or development of a new tumour. The outcomes of the donors and recipients were recorded.Both donors are alive with no clinical or radiological evidence of local, contralateral or distant recurrence of the tumour. Neither of the recipients have developed any suspicious lesions in the transplant kidneys or distant metastasis elsewhere. However, the recipient in the first case developed a haemangiosarcoma in a right brachiocephalic arteriovenous fistula 18 months after the transplant. This required a high above-elbow amputation. Both grafts have been functioning well.Several studies have demonstrated a significant survival advantage with transplantation over dialysis, particularly in elderly patients and in those with co-morbidities.et al reported 43 transplant patients with previously diagnosed small RCC.et al reported no recurrence of the RCC in 3 patients after a median follow-up of 15 months.There are few reports of transplanting kidneys with small RCC. Brook In our study as well as in other studies, the kidney recipients were high risk dialysis patients in whom the morbidity of dialysis outweighed the risks of transplanting kidneys containing a small RCC. Transplantation provides a substantial survival advantage compared with dialysis in this group of patients.Our approach in the above cases was to engage in thorough multidisciplinary team discussion, to explain to both the donors and recipients regarding all the possible consequences and to follow up the donors and recipients closely with ultrasonography, CT and biopsy of any suspicious lesions. We recommend that these kidneys are used only in high risk recipients where the risks of dialysis outweigh the risks of tumour recurrence. There is no need for a change in immunosuppression protocol in these cases.The outcome after kidney transplants following the removal of a small RCC is comparable with that of a normal kidney. However, this should be restricted to high risk recipients in whom the risks of dialysis outweigh the risks of tumour recurrence. Strict follow-up is essential.


The telomere lengths in the paired samples showed a robust relationship between the donor and grafted hepatocytes , demonstrating the feasibility of our Q-FISH method for cell-specific evaluation. While 8 pairs showed no significant difference between the telomere lengths for the donor and the recipient, the other 4 pairs showed significantly shorter telomeres in the recipient than in the donor. Multiple regression analysis revealed that the donors in the latter group were older than those in the former (p = 0.001). Despite the small number of subjects, this pilot study indicates that donor age is a crucial factor affecting telomere length sustainability in hepatocytes after pediatric LDLT, and that the telomeres in grafted livers may be elongated somewhat longer when the grafts are immunologically well controlled.Along with the increasing need for living-donor liver transplantation (LDLT), the issue of organ shortage has become a serious problem. Therefore, the use of organs from elderly donors has been increasing. While the short-term results of LDLT have greatly improved, problems affecting the long-term outcome of transplant patients remain unsolved. Furthermore, since contradictory data have been reported with regard to the relationship between donor age and LT/LDLT outcome, the question of whether the use of elderly donors influences the long-term outcome of a graft after LT/LDLT remains unsettled. To address whether hepatocyte telomere length reflects the outcome of LDLT, we analyzed the telomere lengths of hepatocytes in informative biopsy samples from 12 paired donors and recipients (grafts) of pediatric LDLT more than 5 years after adult-to-child LDLT because of primary biliary atresia, using quantitative fluorescence  These twelve cases were considered suitable for our study protocol, which involved examination of a pair of liver biopsy specimens from both the donor and recipient after a follow-up period of more than 5 years after LDLT. No other selection criteria were adopted. These informative cases were obtained from among 134 recipients who had undergone LDLT at the Jichi Medical University Hospital between May 2001 and March 2009. The overall patient survival rate in our department was 95.0% (134/141). The ages, relationships  and post-LT survivals of the 12 pairs of donors and recipients are listed in The ethics committees of Jichi Medical University and Tokyo Metropolitan Institute of Gerontology approved the use of liver biopsy for this study. The participants provided written informed consent before each liver biopsy in accordance with the Declaration of Helsinki. Parental written permission was obtained when the patients were younger than 20 years old.The surgical procedure (left side liver graft) and postoperative management (immunosuppression started using tacrolimus and steroid) for LDLT were basically the same as described previously Episodes of acute cellular rejection (ACR) are summarized in The liver specimens were obtained by ultrasonography-guided core needle (16 gauge) biopsy from the donor and recipient under local anesthesia. These paired biopsies were performed within a week of each other. The donors fully approved biopsy of their liver because they expressed a desire to know the pathological state of the liver and the pathological relationship between the donor and recipient livers after receiving a full explanation about all the potential complications of the biopsy.The liver biopsy specimens were fixed in 10% buffered formalin solution and embedded in paraffin. The specimens were then sliced into sections 3 μm thick for hematoxylin and eosin (HE) staining and into sections 2 μm thick for Q-FISH. All sections included more than 6 portal tracts. Histological findings were assessed by a histopathologist who was an expert in liver pathology (K.M), and described based on the criteria proposed by the Banff Working Group CCCTAACCCTAACCCTAA peptide nucleic acid probe  and the centromeres were labeled with a FITC-labeled CTTCGTTGGAAACGGGGT peptide nucleic acid probe (CENP1: a non-specific centromere probe) . The chromosome preparations were counterstained with 40, 6-diamino-2-phenylindole . Microscopical images were captured with the Image-Pro Plus software package , and analyzed using our own telomere analysis software ‘TissueTelo Ver. 2’. The telomere length estimate for each nucleus was defined as the ratio of the detected signal intensity of the telomere relative to that of the centromere (telomere-to-centromere ratio: TCR). At least 127 cells  were analyzed in each case.The slides were processed with the Q-FISH method, as reported previously As a control for variations in sample preparation, we also performed Q-FISH on a cell block-section of a cultured fibroblast strain, TIG-1 The main limitation of this study was the small number of subjects for whom NTCR values for the paired donor and graft could be determined. However, despite the limited number (12 pairs) of informative cases, the major findings were statistically significant.p<0.05.The values of measured variables were expressed as mean ± standard deviation or median with a range of values. The median NTCRs of the donor and recipient hepatocytes were compared between cases using the Mann-Whitney test or Wilcoxon test. Correlations were analyzed with the Spearman correlation coefficient test and a single regression analysis using the software package Dr.SPSS II . The relationships among NTCR values, donor age and group (lower versus comparable NTCR) were assessed by multiple regression analyses using StatView software version 5.0 (SAS Institute Inc.). Differences were considered significant at p = 0.0038) (p = 0.092).Scatter plot analysis of the median NTCR values for hepatocytes provided a robust correlation between donors and recipients ( 0.0038) . The slop = 0.017)  . Conversp<0.0001), and that the difference between the groups was significant (p = 0.001)  .p = 0.799). Also, neither the NTCR values for recipients nor those for the donors differed significantly between the two groups .With regard to other factors, the ages of the recipients at LDLT showed no significant difference between the two groups  of the 8 donors in the comparable NTCR group showed moderate steatosis.Among the liver biopsy samples from recipients, significant abnormalities were observed in all of the 4 recipients in the lower NTCR group  and in 4 of the 8 recipients in the comparable NTCR group  .p = 0.056). The recipient in pair 11, who has recently achieved complete withdrawal of immunosuppression, also had a higher median NTCR than the donor. In only one recipient (pair 8) who received a graft from a non-father non-mother (stepfather) donor and was treated with a calcineurin inhibitor, the median NTCR was also higher than that of the donor, but the difference was of borderline statistical significance (p = 0.065). While only one of 8 recipients in the comparable NTCR group (pair 6) continued to require oral maintenance steroid therapy, 2 of 4 recipients (pairs 2 and 3) in the lower NTCR group did so.In the comparable NTCR group, 3 recipients  showed a benign course after LDLT, and were treatable with a calcineurin inhibitor at a frequency of less than once a day, or without any immunosuppressant. In particular, the recipient in pair 10, who achieved complete withdrawal of immunosuppression, had a higher median NTCR than the donor, but the difference was of borderline statistical significance (Episodes of acute cellular rejection (ACR) were observed in 2 cases in the lower NTCR group and 4 cases in the comparable NTCR group. Among the cases associated with ACR, 4  showed decreasing NTCR values in the recipients, and 2  showed increasing NTCR values in the recipients, but not to a significant degree. With regard to steatosis, 3 of the 4 cases in the comparable NTCR group associated with ACR in the recipient showed steatosis in the donor.The laboratory data for recipients  and a lower group (showing a significantly lower NTCR in the recipient than in the donor). However, in 5 of the 8 cases in the comparable group, hepatocyte NTCR values for recipients were larger than those for the donors. Secondly, donors in the lower NTCR group were significantly older than those in the comparable NTCR group, and there was no relationship between the absolute NTCR values With regard to the methodology used for telomere length measurement, numerous approaches have been adopted, including Southern blotting, PCR-based methods, and Q-FISH. Previous studies using Southern blotting have shown that fibrosis or hepatitis can accelerate telomere shortening in the liver Our previous population studies demonstrated that telomeres in the liver shorten most rapidly with age among those in the major organs. The telomere shortening in liver is especially rapid in infants, and then the rate of shortening slows from adolescence to middle age; no significant decrease is evident from forties to centenarian age et al. have reported that non-alcoholic fatty liver disease with steatosis causes prominent telomere shortening in hepatocytes The regeneration of a liver graft in a recipient suggests that the structure and function of the graft might be largely controlled by the host environment With regard to immunological findings, we recently assessed the association of ACR with several factors in 114 LDLTs performed at our institute, including the cases analyzed in the present study. We found that paternal grafts with gender mismatch were associated with a higher incidence of ACR than maternal grafts with gender match, and that there was no significant difference between the donor age groups With regard to the relationship between recipient status and LT/LDLT outcome, some studies have demonstrated a correlation between the PELD score before LT and outcome after LT Major arguments concerning the clinical impact of donor age on LT/LDTP outcome have focused mainly on the elderly (60∼65 years of age or more). However, aging phenomena have been observed not only in the elderly but also in young adult or middle-aged individuals Notwithstanding our retrospective study design and the limited number of samples analyzed, our findings lend additional support to the validity of LDLT, because telomere lengths in the majority of the recipients were statistically comparable to those in the native liver of the donor. Furthermore, telomere lengths showed a tendency to be elongated in well-controlled grafted livers. Our data also provide essential clues to the relationship between hepatocyte aging and LDLT outcome, particularly with regard to young/middle-aged individuals. Further studies will undoubtedly reveal other factors that affect hepatocyte telomere length in the grafted liver and their long-term effects on the graft after pediatric LDLT.Table S1Laboratory data of recipients and donors at biopsy. Lower NTCR group: Median NTCRs of the recipient hepatocyte was significantly lower than those of the donor. Comparable NTCR group: Median NTCRs of the recipient hepatocyte was neither significantly lower nor higher than those of the donor. AST: aspartate aminotransferase, ALT: serum alanine aminotransferase, γ-GTP: γ-glutamyl transpeptidase, Alb: albumin, T-bil: total bilirubin, PT-INR: international normalized ratio of prothrombin time.(DOC)Click here for additional data file.Table S2Pediatric End-stage Liver Disease (PELD) score of patients. Alb: albumin, T-bil: total bilirubin, PT-INR: international normalized ratio of prothrombin time.(XLS)Click here for additional data file.Table S3Acute cellular rejection episodes and Immunosuppression status. ACR: Acute Cellular Rejection.(XLS)Click here for additional data file.


P < 0.05) and peaked rapidly at day 5 after injury, but decreased thereafter. The CSF endostatin concentration in 12 patients with an unfavorable outcome was significantly higher than that in 18 patients with a favorable outcome at day 5 (P = 0.043) and day 7 (P = 0.005) after trauma. Receiver operating characteristic curve analysis suggested a reliable operating point for the 7-day CSF endostatin concentration predicting poor prognosis to be 67.29 pg/mL. Our preliminary findings provide new evidence that endostatin/collagen XVIII concentration in the CSF increases substantially in patients with sTBI. Its dynamic change may have some clinical significance on the judgment of brain injury severity and the assessment of prognosis. This trial is registered with the ClinicalTrials.gov Identifier: NCT01846546.Recent studies have suggested that endogenous angiogenesis inhibitor endostatin/collagen XVIII might play an important role in the secondary brain injury following traumatic brain injury (TBI). In this study, we measured endostatin/collagen XVIII concentrations serially for 1 week after hospitalization by using the enzyme-linked immunosorbent assay method in the cerebrospinal fluid (CSF) of 30 patients with TBI and a Glasgow Coma Scale (GCS) score of 8 or less on admission. There was a significant trend toward increased CSF levels of endostatin after TBI versus control from 72 h after injury. In patients with GCS score of 3–5, CSF endostatin concentration was substantially higher at 72 h after injury than that in patients with GCS score of 6–8 ( It isn injury , nothingBetween October 2006 and March 2007, we conducted a prospective cohort observational study of patients with severe TBI  score of 8 or less) requiring continuous lumbar drainage of CSF in the neurosurgery ward of the Sixth People's Hospital affiliated to Shanghai Jiaotong University. Patients delivered within 4 h whose highest abbreviated injury score (AIS) was 3 or less (other than head injury) were considered to be isolated TBI cases and were included. To avoid interfering factors, patients who suffered open or combined injuries or had existing prior neurological disease were excluded. Those who died within the first week of hospitalization and/or whose serial CSF samples could not be obtained were also excluded as having incomplete data. Accordingly, 30 patients were analyzed in this study. Demographic data, including age, gender, mechanism of trauma, and GCS score at admission, were documented when the patients arrived at the emergency room. The control group comprised 20 patients whose CSF was examined via lumbar puncture for investigation of suspected neurological disease. All patients had normal neurological examination and negative imaging studies, and none had evidence of trauma or preexisting neurological disease, including tumors, vascular anomalies, or abnormalities of CSF. The study had approval from the hospital ethics committee and an informed consent for participating in the study was obtained from an appropriate member of each patient's family before performance of lumbar drainage.Immediately after admission, all patients were evaluated with an initial CT scan and were followed with serial neurological examinations. All the imaging studies were technically adequate and were reviewed by the staff of the radiology department. Subsequently, insertion of a lumbar drain into the subarachnoid space was conducted in a standard fully sterile technique. The lumbar subarachnoid catheter was placed in the subarachnoid place with 15 cm in length via lumbar 4-5 or 3-4 interspinous space through a spinal needle, using routine puncture procedure with the patient in lateral decubitus position. The remaining part of the catheter was positioned to course transversely across the back and slightly up the side of the abdomen. The catheter insertion site was treated with povidone-iodine ointment and the entire catheter was covered with drape. The distal end of the catheter was hooked up to an empty transfer pack with a macrodrip chamber that allows accurate quantification of CSF drained. This system used a standard lumbar drain connected to an intravenous infusion pump to provide drainage of CSF in a constant and predictable manner. The patients were kept at absolute bed rest but allowed to turn from side to side and could not sit up to about 45° angle in bed. The drainage chamber was adjusted with respect to the patients head so that the drainage rate could be altered and overdrainage could be prevented. The desired drainage rate was approximately 5–15 mL/hour or 120–360 mL/day. The system was left in place for 7–10 days. At the end of the course of external drainage, the drain was left closed. The development of any neurological findings should prompt rapid discontinuation of lumbar drainage and immediate radiographic evaluation. There was no evidence of complications, such as tension pneumocephalus, brain herniation, and intracranial infection. All patients were evaluated and treated according to the guidelines of the “Management of Severe Head Injury” in the neurosurgery intensive care unit . OutcomeCSF samples from patients with severe TBI were collected serially from the lumbar drainage system at 1, 3, 5, and 7 d after injury. CSF samples from the controls were also collected from the lumbar subarachnoid space. Samples were centrifuged to remove cellular debris, and the supernatant was immediately frozen at −20°C after sampling and stored for later analysis. y-axis against the concentration on the x-axis and a best fit curve was drawn through the points on the graph. The data may be linearized by plotting the log of the endostatin/collagen XVIII concentrations versus the log of the optical density, and the best fit line can be determined by regression analysis. Consequently the endostatin/collagen XVIII concentrations in CSF samples were obtained by reading form the standard curve.Endostatin/collagen XVIII concentrations in CSF were determined by enzyme-linked immunosorbent assay (ELISA) with commercially available kits , according to the manufacturer's instructions. This assay employs the quantitative sandwich enzyme immunoassay technique. In brief, a monoclonal antibody specific for endostatin/collagen XVIII had been pre-coated onto a microplate. Standards and CSF samples were pipetted into the wells and any endostatin/collagen XVIII present was bound by the immobilized antibody. An enzyme-linked monoclonal antibody specific for endostatin/collagen XVIII was added to the wells. Following a wash to remove any unbound antibody-enzyme reagent, a substhumane solution was added to the wells and color developed in proportion to the amount of endostatin/collagen XVIII bound in the initial step. After addition of stop solution into each well, the color development was stopped and the optical density was measured within 30 minutes, using a microplate reader set to 450 nm. No significant cross-reactivity or interference was observed in this assay. Finally, a standard curve was constructed by plotting the mean absorbance for each standard on the t-test was used for comparisons between TBI and control patients. The CSF concentrations were not normally distributed; therefore, group means were compared using the Mann-Whitney test. Fisher's exact test was used to compare proportions where appropriate. Statistical significance was assumed for P values of less than 0.05. A receiver operating characteristic (ROC) curve was used to assess the accuracy of predictions of poor outcome from the CSF endostatin/collagen XVIII concentrations. Using the ROC curve, a CSF endostatin/collagen XVIII concentrations cutoff value and its confidence in prognosis could be estimated based on the area under the ROC curve (AUC).The statistical package Statistical Program for Social Sciences  was used for analyses. All values are expressed as mean ± standard error of the mean (SEM) unless otherwise specified. A two-tailed Student's Of the 30 patients with severe TBI, twenty were male and ten were female. The patients' age ranged from 16 to 82 years, with mean age of 51.1 years. The mechanisms of trauma included motor vehicle collisions, falls, heavy strikes , and assaults. Types of lesions, as evidenced by radiologic and neurologic symptoms or signs, included cerebral contusions/lacerations, intracranial hematomas, brain stem injury, and diffuse axonal injury. The control group contained thirteen male and seven female. Their age averaged 52.1 years and ranged from 34 to 66 years. Clinical characteristics of the two groups are summarized in P > 0.05), its levels in CSF rapidly increased thereafter (P < 0.05) and reached almost twice that of controls at day 5  and day 7   .P > 0.05) (P < 0.05), and it peaked rapidly at a mean value of 87.47 ± 16.32 pg/mL at day 5 after injury, approximately 1.5 times that in patients with GCS 6–8  but declined to 54.09 ± 9.09 pg/mL 7 days after trauma, not significantly lower compared with the GCS 6–8 group at day 7    and extra severe TBI (GCS 3–5)  groups. These two groups did not differ significantly in age or sex ratio, nor was a significant difference observed in the mean CSF endostatin/collagen XVIII concentration at 24 h after injury between the two groups  . In pati > 0.05) .P = 0.026) and day 5 (P = 0.001), although this did not reach statistical significance at 24 h and 7 days after trauma . Death group showed a trend toward increased CSF concentrations of endostatin/collagen XVIII after TBI versus survival group, which was especially significant at day 3 (r trauma .P = 0.043) and day 7 after trauma (P = 0.005) (P = 0.316) and 72 h (P = 0.230) after injury .At 6 months after TBI, outcome was favorable for 18 patients  but unfavorable for twelve patients . No significant difference was found between these two groups in age or sex ratio. The CSF endostatin/collagen XVIII concentration was significantly higher in patients with an unfavorable outcome than in patients with a favorable outcome at day 5 (= 0.005) . No diffr injury . Using tr injury ; the AUCRevascularization therapy to enhance trophic blood supply is now considered promising for the treatment of patients who suffered TBI, although angiogenesis may be of importance for the secondary brain damages development –3. DurinPrevious clinical and experimental pieces of evidence have proved that endostatin/collagen XVIII was prevailingly expressed and secreted during the early activation of microglia/macrophages after TBI –12. Dein+ cells are not only accumulated at the lesion site, in pannecrotic debris zone, and perivascular Virchow-Robin spaces (the drainage route for blood-borne leucocytes), but also at the lesion margin after TBI and spinal cord injury [Endostatin/collagen XVIII has been shown to have a more profound affect on tumor neovasculature. It plays certain inhibiting roles in axon sprouting and neuronal and glial development, although the mechanism of its actions remains undefined. However, the role of endostatin during the tissue remodelling response to CNS injury remains largely elusive. Mueller et al. and Zhang et al. observed that endostatin/collagen XVIIId injury , 11, 18.2O2 and hypoxia and modulated by nitric oxide pathway [+ vessels matched with the districts in which VEGF expression was described by others earlier extending up to approximately 30–100 μm. Shore et al. detected a peak of VEGF levels in CSF of infants and children at 22.4 hours after TBI. But we have not investigated the peak of CSF endostatin/collagen XVIII levels till the end of the observation period. Further investigation is warranted to determine whether this increase is associated with an increase in CSF VEGF and determine the role of VEGF in triggering the increase in CSF endostatin/collagen XVIII concentration.As potential mediator of vascularization, the proangiogenic factor VEGF is upregulated at the lesion site during the period of maximal endothelial proliferation following TBI , giving  pathway . Accordi pathway , 5, 8, DThere are several limitations to our study. First, it is unclear whether CSF endostatin/collagen XVIII concentration from the lumbar subarachnoid space reflects simultaneous brain intracellular or interstitial concentration. Second, the number of patients is small and selectively limits the power to detect potential effects due to differences in endostatin/collagen XVIII concentration, injury severity, and clinical outcome. Third, our analysis is restricted to CSF samples taken from the patients requiring lumbar drainage of CSF. This might have introduced a significant bias in patient selection, and it is therefore difficult to confirm the relationship between the type of TBI and CSF levels of endostatin/collagen XVIII. In addition, the concentration of endostatin/collagen XVIII in the CSF of control group may not represent the normal baseline because of the interference caused by the controls' potential disease of other system. Finally, there were minor variations in the treatment of the TBI patients, although these patients were all treated by the same neurointensive care team using the same standardized protocol.We describe the time-dependent increase of endostatin/collagen XVIII levels in CSF of patients with severe TBI that elevated within the first week after injury. This observation supports the hypothesis that endostatin/collagen XVIII plays an important role in the response of the CNS to injury, possibly involving development of secondary brain damage. However, we could not uncover its role in this process. It may provide a theoretical basis for a new pharmacotherapy, selectively attenuating the endostatin/collagen XVIII expression by NO synthase inhibitor, to intervene secondary brain damages. Further studies are also needed to clarify the precise relationship between CSF VEGF and CSF endostatin/collagen XVIII in severe TBI.


These neurons were shown recently to express a polycystin member of the TRP channels family (PKD2L1) that potentially acts as a chemo- or mechanoreceptor. Recent studies carried out in young rodents indicate that spinal CSF-cNs express immature neuronal markers that appear to persist even in adult cells. Nevertheless, little is known about the phenotype and morphological properties of medullar CSF-cNs. Using immunohistochemistry and confocal microscopy techniques on tissues obtained from three-month old PKD2L1:EGFP transgenic mice, we analyzed the morphology, distribution, localization and phenotype of PKD2L1+ CSF-cNs around the brainstem and cervical spinal cord central canal. We show that PKD2L1+ CSF-cNs are GABAergic neurons with a subependymal localization, projecting a dendrite towards the central canal and an axon-like process running through the parenchyma. These neurons display a primary cilium on the soma and the dendritic process appears to bear ciliary-like structures in contact with the CSF. PKD2L1+ CSF-cNs present a conserved morphology along the length of the medullospinal central canal with a change in their density, localization and dendritic length according to the rostro-caudal axis. At adult stages, PKD2L1+ medullar CSF-cNs appear to remain in an intermediate state of maturation since they still exhibit characteristics of neuronal immaturity  along with the expression of a marker representative of neuronal maturation (NeuN). In addition, PKD2L1+ CSF-cNs express Nkx6.1, a homeodomain protein that enables the differentiation of ventral progenitors into somatic motoneurons and interneurons. The present study provides valuable information on the cellular properties of this peculiar neuronal population that will be crucial for understanding the physiological role of CSF-cNs in mammals and their link with the stem cells contained in the region surrounding the medullospinal central canal.The mammalian spinal cord and  Some of these organs form a bridge between the CSF, the bloodstream, and neurons in the parenchyma and are also believed to be involved in neuronal volume transmission In mammals, most of our knowledge concerning the medullospinal CSF-cN system comes from studies conducted at the spinal cord level + CSF-cNs are also present in the dorsal vagal complex (DVC), a hindbrain region dedicated to the regulation of important autonomic functions + and sensitive to changes in pH and osmolarity of the extracellular fluid. These neurons displayed robust action potential discharge activity as well as GABA and glycinergic synaptic currents. However, their phenotype and their degree of maturity have not yet been determined.The chemical hypothesis is supported by the presence of various bioactive molecules such as hormones and neurotransmitters circulating in the CSF that are thought to come in contact with CSF-cNs A renewed interest in these particular neurons emerged from the demonstration that the region surrounding the spinal cc, where CSF-cNs are found, represents a stem cell niche and may generate both neurons and/or glial cells in lower vertebrates + CSF-cNs in the cervical spinal cord and brainstem of adult mice. Secondly, we sought to quantitatively characterize the degree of cellular maturity of these cells by measuring markers of immature and mature neurons, and by doing so to additionally compare the phenotype of medullar S-CSF-cNs to that of cervical spinal CSF-cNs.Our first aim was therefore to carry out an in-depth analysis of the morphology, distribution and localization of PKD2L1We show that S-CSF-cNs are present, with a conserved morphology, along the entire length of the medullospinal cc and bear a primary cilium on the soma, not on the bud. These cells are mainly GABAergic and strongly express PKD2L1 proteins on the somatodendritic compartment. Our findings confirm that this channel represents a specific marker for medullospinal CSF-cNs. In addition, we demonstrate that, at adult stages, CSF-cNs remain in an intermediate stage of maturity, as they continue to express DCX, Hu/CD and Nkx6.1 in the presence of NeuN.ad libitum.This study was carried out in strict conformity with the recommendations and rules set by the EC Council Directive (2010/63/UE) and the French “Direction Départementale de la Protection des Populations des Bouches-du-Rhône”. Prior to any experimental procedure, animals were anaesthetized using a mixture of Xylazine/Ketamine and euthanized (see below). All efforts were made to insure animal well-being and minimize animal suffering and the number of animals used. The experimental procedures have been approved by our local Animal Care Ethical Committee (Comité Ethique de Provence N°14) (licence N°13.435 held by JT and N°13.430 by NW). Animals were housed at constant temperature (21°C) under a standard 12 h light-12 h dark cycle, with food  and water provided + cells, EGFP expression was selectively induced by CRE recombinase activity + animals, we carried out PCR on tail genomic DNA. CRE transgene was detected using the forward primer 5′-CGT ACT GAC GGT GGG AGA AT-3′ and the reverse primer 5′-CCC GGC AAA ACA GGT AGT TA-3′, with the following PCR conditions: 7 min at 95°C, 35 cycles at 95°C for 40 sec, 62°C for 40 sec and 72°C for 40 sec, followed by a final step at 72°C for 6 min. The amplicon size was 166 bp. EGFP was detected using the forward primer 5′-GCC ACA AGT TCA GCG TGT CC-3′ and the reverse primer 5′-GCT TCT CGT TGG GGT CTT TGC-3′, with the following PCR conditions: 5 min at 95°C, 36 cycles at 95°C for 30 sec, 64°C for 30 sec and 72°C for 40 sec, followed by a final step at 72°C for 7 min. The amplicon size was 573 bp.We used PKD2L1:EGFP positive and negative transgenic mice obtained by crossing PKD2L1-IRES-Cre with Z/EG reporter transgenic mice. Thus, in PKD2L1Three-month-old adult mice were anaesthetized with an intraperitoneal injection of ketamine  and xylazine  mixture  and transcardially perfused with phosphate buffer solution (PBS at 0.1 M). Subsequently the animals were perfused with 4% PFA in PBS. For the experiments where GABA immunoreactivity was tested, PKD2L1:EGFP positive animals were transcardially perfused with a fixation medium containing 4% PFA and 0.2% glutaraldehyde. Brains and spinal cords were immediately removed, post-fixed one hour in 4% PFA at 4°C, rinsed in PBS , cryoprotected for 24 to 48 hours in 30% sucrose at 4°C and frozen in isopentane (−40°C).Brainstem and cervical spinal cord coronal or sagittal thin sections (40 µm) were obtained using a cryostat (Leika CM3050) and collected serially in twelve-well plates containing 0.1 M PBS. In the analysis carried out in our study, we distinguished the cervical spinal cord region  are represented with white dashed lines. Low resolution images of full sections  were also acquired to allow identification of the section’s position in the rostro-caudal axis using specific anatomical landmarks and to orient each section in the dorsal-ventral axis.Sections were observed on a confocal laser scanning microscope  equipped with solid state fiber optic lasers and single plane images or stacks of images were acquired with a 2x digital zoom using either a 20x objective  or a 63x oil immersion objective . Images have been acquired at optimal resolution between 512×512 and 1024×1024 pixels. To insure the continuity of the image acquisition, each plane acquired for a stack overlapped with the next plane. When using two fluorochromes with different excitation/emission spectra, images were acquired sequentially for each channel  and the filters and photomultiplier tubes settings chosen to optimize images and avoid signal crosstalk. The prepared images were obtained from a maximum intensity Z-projection. In all images illustrating coronal or sagittal sections, the limits of the central canal (cc) or 4All analyses, cell counting and determination of markers co-expression were carried out on single images from stacks with cell to cell identification. The analyses were performed on non consecutive sections. Images were analyzed and prepared using ZEN 2009 light Edition (Zeiss software), ImageJ 1.45 (NIH), Adobe Illustrator and Photoshop. No correction was applied to the images and for a better visualization, especially for thin structures like fibers, only contrast and brightness were adjusted in the images used for the figures. When images of a same set of experiments are illustrated, the same adjustments were applied to allow comparison.+ cells were counted manually in every image within the corresponding image stack (14 to 24 images/stack and 15 to 26 µm stack depth) using the ‘cell counter’ ImageJ plug-in. Subsequently, the total number of cells counted was divided by the thickness of the analyzed stack and cell density expressed per 10 µm of tissue depth. To analyze the distribution of PKD2L1+ cells around the cc, each stack of images was oriented in the dorsal-ventral axis using the corresponding low resolution image. The images were then subdivided in 4 quarters centered on the cc as illustrated in + S-CSF-cNs and the border of the cc (straight line). To determine the neurite length of S-CSF-cNs depending on their position in a specific quarter and a specific level (see above for details), a segmented line (ImageJ) was used to follow over several optical slices the path of the neurite from the soma to the bud.Cell density, distribution and morphology. At each medullospinal level of interest , we manually counted the number of cells expressing PKD2L1/EGFP and the number of cells expressing the marker of interest. Subsequently, we calculated the percentage of PKD2L1+ cells co-expressing both markers. In experiments where the investigated marker was nuclear (NeuN except Nkx6.1), we first created a ROI at the size of a S-CSF-cN nucleus using ImageJ. This ROI was then used to measure in each image of a stack both the background average intensity  in a region without cell bodies or fibers  and the nuclear average intensity in all PKD2L1+ cells . Out of the values obtained for each image in a determined stack, we then calculated the mean background intensity and only PKD2L1+ cells with an average nuclear labeling intensity greater than 30% of the background value were considered positive for NeuN.Cell counting in double labeling experiments. In experiments where we quantified the co-expression of selected cytoplasmic markers in PKD2L1All data are expressed as mean ± S.E.M and the statistical significance was tested using a Kruskal-Wallis test (non-parametric) combined with a Dunn’s multiple comparison tests. Data were considered to be significantly different given a P value below 0.05. Analyses were carried out using Excel 2007 (Microsoft) and Prism (Graphpad).+ S-CSF-cNs along the central canal. Using an immunohistochemical approach with primary antibodies directed against PKD2L1, we first specified the morphology of PKD2L1+ CSF-cNs and compared it along the length of the medullospinal cc on coronal sections from cervical spinal cord and brainstem. + cells can be observed around the cc. Typically, the large majority of PKD2L1+cells  presented the classical morphology described in mammals for CSF-cNs around the cc (fig. A1 arrowheads and B) + cells had a small soma with an average diameter of 8.0±0.1 µm (336 cells) and projected a thick neurite (see below) to the cc that ended with a protrusion or bud . We further noticed that PKD2L1+ CSF-cNs regularly exhibited an enlargement along the thick neurite  and through the brainstem  to 4 . Indeed, when counting PKD2L1+ CSF-cNs per 10 µm of tissue depth on images obtained from coronal sections, we found: 10±1 cells at level 1, 8±1 and 7±1 at levels 2 and 3, respectively and only 3±1 at level 4  and dorsal (on average 33±2%) position with less cells present laterally  and in parallel the neurite length of lateral S-CSF-cNs increased to 32.5±2.3 µm (n = 22) at level 4  along the rostro-caudal axis, we noticed that PKD2L1+ S-CSF-cNs and identified the somatodendritic versus axonal compartments using subcompartment-specific antibodies  revealing their classical morphology but also numerous ciliary structures around the cc (+ S-CSF-cNs suggested the presence of Ac-αTub+ ciliary-like structures on their bud (+ mice to determine whether the structures observed on the bud were primary cilia (+ S-CSF-cNs exhibiting AC3 labeled cilia on their soma (White open arrowheads) but not on the bud (White arrowheads). We observed these primary cilia on the soma of all PKD2L1:EGFP+ S-CSF-cNs with an average length of 3.4±0.2 µm (n = 153) and a diameter in the sub-micrometer scale.Medullar S-CSF-cNs possess a primary cilia on their soma. In spinal cord, the presence of cilia on the bud of CSF-cNs has been reported in several vertebrate species d the cc . Interesd the cc a and c. heir bud  and 4. Sry cilia . We did ry cilia . Figure + CSF-cNs are present along the length of the cc with an exclusive subependymal position (S-CSF-cNs). They exhibit a conserved morphology with a dendrite projecting to the cc that ends with a protrusion that appears to present a ciliary structure; they also exhibit a primary cilium on their soma. Furthermore, these cells strongly express PKD2L1 only on their somatodendritic compartment and this protein appeared to be a specific maker for this cell population. Finally, the analysis of their distribution from spinal cord to brainstem indicates that their density decreases and that both their position around the cc and the length of their dendrite changes along the rostro-caudal axis.Taken together, the results presented above indicated that PKD2L1+ mice. The selectivity of the GABA/GAD67 immunolabeling was confirmed by the presence of GABA+/GAD67+ cells in brain regions known to contain GABAergic neurons: the spinal cord dorsal horn, the nucleus tractus solitari and the cerebellum (Data not shown). As visualized in + S-CSF-cNs were also GABA+ .Earlier studies have indicated that spinal CSF-cNs were GABAergic in rodents + medullar S-CSF-cNs coexpressed GAD67  and that overall 79±2% of S-CSF-cNs were PKD2L1+/GAD67+ (level 1–4∶577 GAD+ cells out of 733 PKD2L1+ S-CSF-cNs). In these cells we primarily observed GAD67+ staining delineating the soma and the bud of PKD2L1+ S-CSF-cNs , we found that on average 66±3% of PKD2L1+ S-CSF-cNs expressed NeuN . Further, our results indicated that the proportion of PKD2L1+ S-CSF-cNs expressing NeuN progressively increased from 54±4% at level 1 to 100% in the most rostral region of the cc . The entire population of PKD2L1+ S-CSF-cNs from the cervical spinal cord up to the rostral part of the brainstem also strongly expressed Nkx6.1 . Further, using a selective antibody against DCX, we also demonstrated that 90±1% of medullar PKD2L1+ S-CSF-cNs (level 2) coexpressed DCX  expressed HuC/D . Finally, although the number of PKD2L1+ S-CSF-cNs expressing DCX appeared similar at all 4 levels, there was a tendency for a decrease in the number of S-CSF-cNs that expressed HuC/D along the rostro-caudal axis  and the brainstem (level 2–4) in adult animals. Furthermore, we carried out a quantitative and comparative analysis of protein expression levels within these cells.+ cells around the cc: one predominant population with the previously reported morphology for CSF-cNs + cells in the medullospinal ependymal layer of mice tissue, indicating the absence of intraependymal CSF-cNs. Secondly, we observed that all MAP2+ CSF-cNs coexpressed PKD2L1. We therefore concluded that, in mice, PKD2L1 represents a selective marker for medullospinal CSF-cNs and that these neurons have an exclusively subependymal localization.Using an immunohistochemical approach with antibodies against PKD2L1, we observed two groups of PKD2L1+ and NF-160 negative) through the ependymal layer. This dendrite ends in the CSF cavity with a bud of conserved diameter (∼3 µm) on top of which, we observed, in some instances, a cilia-like structure (see below). Our data are in agreement with previous reports obtained in the mouse spinal cord We showed that along the length of the medullospinal cc, these cells represent a homogenous neuronal population exhibiting a simple morphology with a small round soma (∼10 µm) extending a single thick dendrite  around the cc. We found the largest density at the cervical level of the spinal cord with a progressive decrease as we moved rostrally; fewer PKD2L1+ S-CSF-cNs were observed around the 4V. We also found that the localization of PKD2L1+ S-CSF-cNs around the cc changed from a predominantly dorsal-ventral localization at the cervical spinal cord level to an almost exclusively lateral one at more rostral levels. Moreover, we showed that the dendritic length of S-CSF-cNs also changed along the medullospinal axis in an orientation-dependent manner. At the level of the cervical spinal cord S-CSF-cNs with a ventral localization had the longest dendrites while in the brainstem S-CSF-cNs with lateral localizations had the longer dendrites. Although we do not have, so far, any data regarding the functional consequences of such an organization, one might suggest that this specific distribution and organization along the cc could lead to specific projection pathways as well as innervations of specific territories. Answering this question might give us insights into the specific function of S-CSF-cNs at different levels of the medullospinal cc.We demonstrated that PKD2L1via sodium action potentials to distal partners.PKD2L1 is a member of the polycystin TRP channel subfamily and was first described in the kidney Generally, one distinguishes motile from non-motile/primary cilia on the basis of protein composition and organization + cilia-like structures of a few micrometers of length (1–3 µm) emerging from the bud. Nevertheless, when testing for AC3 immunoreactivity, a selective marker of primary cilia + structures in our sections but in S-CSF-cNs AC3+ cilia where present only on their somata. This is the first demonstration of a primary cilium on the soma of identified medullospinal S-CSF-cNs, and fits well with one recent study using transmission and scanning electron microscopy in mouse spinal cord. There, cells around the spinal cord cc with a morphology of CSF-cNs exhibited on their soma a single cilium of indeterminate identity + S-CSF-cNs would be capable of sensing modifications in the motion or composition of interstitial medium. Interestingly, the presence of polycystin-2, a protein of the same family as PKD2L1, in the basal region of the primary cilium has been described in kidney epithelia and vascular endothelial cells and shown to confer to these cells a role in mechanodetection of the flow of primary urine or blood, respectively + and AC3 negative cilia-like structures on the PKD2L1+ terminal protrusion; a result that might further support its role as a CSF sensor.In the present study, using classical immunohistochemical techniques with antibodies against cytosolic EGFP, we were able to detect some Ac-αTub+ S-CSF-cNs. However, we cannot exclude the possibility that some GABAergic S-CSF-cNs could not be detected due to low levels of GABA/GAD expression. On the other hand, these S-CSF-cNs might represent a neuronal population with a different phenotype as suggested by several studies + S-CSF-cNs could express either the aromatic L-amino acid decarboxylase (AADC) to synthesize monoamines Numerous studies carried out on the spinal cord of several low vertebrates species + staining appeared punctate, in close apposition with the soma of PKD2L1+ S-CSF-cNs and might correspond to GABAergic terminals known to accumulate GAD + S-CSF-cNs are modulated by functional GABAergic terminals likely arising from local interneurons. We also observed fibers positive for TH, 5-HT and ChAT in close contact with PKD2L1+ S-CSF-cNs suggesting some catecholaminergic, serotoninergic or cholinergic modulation of S-CSF-cNs. This is supported on the one hand by the presence of cholinergic neurons in the dorsal motor nucleus of the vagus nerve that could project towards S-CSF-cNs and on the other hand by immunohistochemical studies showing that cells around the cc with the morphology of CSF-cNs were strongly positive for type 6 5-HT receptors www.Gensat.org BAC Address: RP23-65B23). Our results support the idea that S-CSF-cNs are primarily GABAergic cells, receive GABAergic inputs, and might be modulated by other neurotransmitters such as acetylcholine and 5-HT.Complementing these results, we found a strong labeling against GAD67 in the parenchyma and around the cc . This GAin vivo of dividing and generating astrocytes and oligodendrocytes while in vitro they form neurospheres Most CNS neurons go through a specified maturation process, but in regions such as the olfactory bulb or the dentate gyrus of the hippocampus new neurons are still formed even in adulthood + S-CSF-cNs have conserved immature characteristics or are young neurons. Based on several studies indicating that spinal stem cells do not generate new neurons in adults + S-CSF-cNs against PSA-NCAM. This is in contrast to previous reports First, we show that in adult animals the vast majority of S-CSF-cNs still expressed DCX (∼90% of cells) as well as HuC/D although in a lower proportion of cells (∼60% of cells). Since these markers are found both in young post-mitotic and immature neurons and in new neurons formed during adult neurogenesis + S-CSF-cNs strongly expressed Nkx6.1 and second that this high level of Nkx6.1 expression was present in PKD2L1+ S-CSF-cNs from the cervical spinal cord up to the rostral part of the brainstem. We observed little expression in other cells around the cc , ventral (V)2 and V3 neurons d the cc . These r+, PSA-NCAM+ and HuC/D+ phenotype and confirms the immature state of spinal CSF-cNs. These authors also suggested that cells present in the spinal ependyma, presumably CSF-cNs, were negative for NeuN in tissues obtained from 3 week-old rats (P21). In our study however, we showed that in adult mice (∼P90) S-CSF-cNs did express NeuN along with the immature markers (see also +/PKD2L1+ S-CSF-cNs changes along the rostro-caudal axis. In a sense, these results are unexpected since during development or neurogenesis NeuN expression is supposed to increase with neuronal maturation while DCX expression should decrease and disappear in parallel In young rats, Marichal and colleagues see also . Our res+ CSF-cNs in the brainstem of mammals. We show that CSF-cNs correspond to a morphologically homogeneous neuronal population that has a subependymal localization and extends from the cervical spinal cord up to the rostral brainstem. We also demonstrate that S-CSF-cNs exhibit a primary cilium on their soma contacting the interstitial liquid and a protrusion at the end of their dendrite in contact with the CSF. These two appendices might enable S-CSF-cNs to sense both the pericellular fluid and the CSF. We further demonstrate that PKD2L1, selectively expressed on the somatodendritic compartment, represents a specific marker for S-CSF-cNs. Due to modulation of this channel by pH and osmolarity + S-CSF-cNs exist in an intermediate stage of maturity. These neurons display many mature properties: they are polarized cells with an axon-like projection, display a robust action potential discharge activity, are integrated in a neuronal network Our study represents the first in-depth analysis of the morphology and distribution of PKD2L1