Quantitative Metaproteomics and Activity-based Protein Profiling of Patient Fecal Microbiome Identifies Host and Microbial Serine-type Endopeptidase Activity Associated With Ulcerative Colitis.

The gut microbiota plays an important yet incompletely understood role in the induction and propagation of ulcerative colitis (UC). Organism-level efforts to identify UC-associated microbes have revealed the importance of community structure, but less is known about the molecular effectors of disease. We performed 16S rRNA gene sequencing in parallel with label-free data-dependent LC-MS/MS proteomics to characterize the stool microbiomes of healthy (n = 8) and UC (n = 10) patients. Comparisons of taxonomic composition between techniques revealed major differences in community structure partially attributable to the additional detection of host, fungal, viral, and food peptides by metaproteomics. Differential expression analysis of metaproteomic data identified 176 significantly enriched protein groups between healthy and UC patients. Gene ontology analysis revealed several enriched functions with serine-type endopeptidase activity overrepresented in UC patients. Using a biotinylated fluorophosphonate probe and streptavidin-based enrichment, we show that serine endopeptidases are active in patient fecal samples and that additional putative serine hydrolases are detectable by this approach compared with unenriched profiling. Finally, as metaproteomic databases expand, they are expected to asymptotically approach completeness. Using ComPIL and de novo peptide sequencing, we estimate the size of the probable peptide space unidentified ("dark peptidome") by our large database approach to establish a rough benchmark for database sufficiency. Despite high variability inherent in patient samples, our analysis yielded a catalog of differentially enriched proteins between healthy and UC fecal proteomes. This catalog provides a clinically relevant jumping-off point for further molecular-level studies aimed at identifying the microbial underpinnings of UC.

Pediatric spinal instrumentation.

This article reviews the evolution of spinal instrumentation in the pediatric age group, starting with the cervical spine and atlantoaxial area and ending with the lower spine. The congenital and the acquired conditions which require instrumentation are described. The technical details regarding pediatric instrumentation are alluded to, and finally an attempt is made to predict the future of spinal instrumentation in this age group.

Proteome-wide covalent ligand discovery in native biological systems.

Small molecules are powerful tools for investigating protein function and can serve as leads for new therapeutics. Most human proteins, however, lack small-molecule ligands, and entire protein classes are considered 'undruggable'. Fragment-based ligand discovery can identify small-molecule probes for proteins that have proven difficult to target using high-throughput screening of complex compound libraries. Although reversibly binding ligands are commonly pursued, covalent fragments provide an alternative route to small-molecule probes, including those that can access regions of proteins that are difficult to target through binding affinity alone. Here we report a quantitative analysis of cysteine-reactive small-molecule fragments screened against thousands of proteins in human proteomes and cells. Covalent ligands were identified for >700 cysteines found in both druggable proteins and proteins deficient in chemical probes, including transcription factors, adaptor/scaffolding proteins, and uncharacterized proteins. Among the atypical ligand-protein interactions discovered were compounds that react preferentially with pro- (inactive) caspases. We used these ligands to distinguish extrinsic apoptosis pathways in human cell lines versus primary human T cells, showing that the former is largely mediated by caspase-8 while the latter depends on both caspase-8 and -10. Fragment-based covalent ligand discovery provides a greatly expanded portrait of the ligandable proteome and furnishes compounds that can illuminate protein functions in native biological systems.

Recurrent thoracic intramedullary schwannoma: report of two cases with long term follow up.

Occurrence of intramedullary schwannomas is rare as Schwann cells are generally present in the nerve root and not in the spinal cord. To date, approximately around 60 cases of intramedullary schwannoma in patients without neurofibromatosis have been reported in the English literature, of which only eight were children. We describe two cases of thoracic intramedullary schwannomas that recurred after primary excision. A 10 years old boy presented with weakness of both lower limbs. Magnetic resonance imaging showed a D10 to D12 intramedullary lesion, which was excised near totally and confirmed to be a schwannoma on histopathological examination. The tumour recurred twice after that and was re operated both times. Another 57 years old lady presented with weakness of both lower limbs and a history of being operated in the past for D9-D10 intramedullary lesion. She was re-operated with total removal of the lesion confirmed to be schwannoma on histopathological examination. Intramedullary schwannoma may recur after resection.

Outcome in 32 cases of tethered cord in adults-is intervention justified?

INTRODUCTION: Adults rarely present with tethered cord syndrome, and this review examines whether it is justifiable to perform surgical intervention in this group. METHODS: Between 2003 and 2017, we performed surgical intervention in 32 adults with tethered cord syndrome. The age range varied from 22 to 65 years. Twenty-six had pain, 20 had motor deficits, and 18 had sensory deficits, whereas 17 presented with sphincter disturbances. Three had undergone surgery for meningomyelocele as infants. Six patients had major spinal deformity in the shape of kyphosis or scoliosis. All patients underwent digital radiographs and MR scans of the whole spine. The oldest with a leaking open meningocele was 41 years old. RESULTS: The mean follow-up was 3.2 years. Twenty-one out of 26 patients presenting with pain had their VAS scores improve significantly; 14 had motor deficits which improved. However, sensory deficits got better in only 7 out of 18 patients, and sphincter improvement was documented (via urodynamics studies and bladder ultrasound) in only 4 out of 17 patients. In 15 cases, surgery was performed under neurophysiological monitoring, and overall improvement was documented in 11 of these patients compared with 7 of the remaining 17 patients. Surgery for spinal deformity was performed in 6 patients, and deformity correction as well as pain reduction was achieved in all. CONCLUSION: Surgery leads to significant reduction of pain by untethering and in those with spinal deformity by correction of the same. Improvement in sensory changes and sphincter problems occurred in few patients. Neuromonitoring certainly has improved our results.

Amniotic band syndrome associated with limited dorsal myeloschisis: a case report of an unusual case and review of the literature.

Amniotic band syndrome (ABS) has been known since ancient times. Descriptions in modern medicine have occurred since the mid-nineteenth century. The association of the amniotic band syndrome with tethered cord is rare. It was generally thought to be incompatible with life. Of late, with better imaging, there have been case reports of amniotic bands causing cord tethering in neurosurgical literature, but its association with limited dorsal myeloschisis has hitherto not been described. We report a case of amniotic band syndrome (ABS) associated with a cervical limited dorsal myeloschisis (LDM) in a child of 2 and a half years old. The management of LDM in ABS is essentially the same as in isolated cases.

Ventriculo-subgaleal shunts-broadening the horizons: an institutional experience.

OBJECTIVE: Ventriculo-subgaleal shunt is an established treatment of hydrocephalus following germinal matrix haemorrhage in low birth weight neonates. It is also used in treatment of post-infective hydrocephalus in children. We intend to emphasise the impact of its extended use in multiple clinical conditions to reduce the number of permanent shunt implantation in infants. METHOD: Retrospective review of clinical cases in a single institution from medical records. RESULTS: VSG shunts with low-pressure valve system were useful in variety of hydrocephalus in infants (post-haemorrhagic, post-infective, post-myelomeningocele, post-shunt block, post-traumatic, hydrocephalus associated with brain tumours). A significant number of infants especially those with post-haemorrhagic and post-myelomeningocele hydrocephalus could be made free of permanent shunt placement. CONCLUSIONS: Ventriculo-subgaleal shunt is an effective, less risky temporary solution of hydrocephalus in infants and can be used in a variety of hydrocephalus in children and helps in avoiding shunt dependency in some of them.

Reagent-Controlled Reversal of Regioselectivity in Nucleophilic Fluorination of Spiro-epoxyoxindole: Synthesis of 3-Fluoro-3-hydroxymethyloxindole and 3-Aryl-3-fluoromethyloxindole.

A convenient and metal/catalyst-free approach for the reversal of regioselectivity in nucleophilic fluorination of a wide range of spiro-epoxyoxindoles has been reported simply by altering the nucleophilic fluoride reagents. Py·(HF) x-mediated fluorination at the tertiary sp3-C center of spiro-epoxyoxindole furnishes 3-fluoro-3-hydroxymethyloxindoles, whereas TBAF-mediated fluoride addition at the primary sp3-C center renders 3-fluoromethyl-3-hydroxyoxindoles, which have been utilized for the synthesis of 3-aryl-3-fluoromethyloxindole.

Chiari malformation and atlantoaxial instability: problems of co-existence.

BACKGROUND: Association of Chiari malformation and atlantoaxial subluxation varies. There is a complex relationship between the two, bony and soft tissue pathologies. METHODS: This is a review of various articles available from the literature on the management of Chiari and its association with atlantoaxial instability. RESULTS: We have an experience of operating on 86 cases of paediatric atlantoaxial subluxation, of which 12 had Chiari malformation diagnosed preoperatively (13.95%). Of the 76 children with Chiari malformations operated on by us, 11 had associated atlantoaxial subluxation diagnosed on imaging (14.47%). CONCLUSIONS: Re-alignment and reduction with fixation may be effective in achieving decompression in cases where reduction is possible from posterior approach. In these cases, posterior fixation is all that is required. If reduction is not possible from posterior and there is "fixed" ventral compression, anterior decompression needs to be combined with posterior fixation. In most cases, direct posterior decompression is warranted.

Management and prognosis of acute traumatic cervical central cord syndrome: systematic review and Spinal Cord Society-Spine Trauma Study Group position statement.

PURPOSE: Spinal Cord Society (SCS) and Spine Trauma Study Group (STSG) established a panel tasked with reviewing management and prognosis of acute traumatic cervical central cord syndrome (ATCCS) and recommend a consensus statement for its management. METHODS: A systematic review was performed according to the PRISMA 2009 guidelines. Delphi method was used to identify key research questions and achieve consensus. PubMed, Scopus and Google Scholar were searched for corresponding keywords. The initial search retrieved 770 articles of which 37 articles dealing with management, timing of surgery, complications or prognosis of ATCCS were identified. The literature review and draft position statements were compiled and circulated to panel members. The draft was modified incorporating relevant suggestions to reach consensus. RESULTS: Out of 37 studies, 15 were regarding management strategy, ten regarding timing of surgery and 12 regarding prognosis of ATCCS. CONCLUSION: There is reasonable evidence that patients with ATCCS secondary to vertebral fracture, dislocation, traumatic disc herniation or instability have better outcomes with early surgery (< 24 h). In patients of ATCCS secondary to extension injury in stenotic cervical canal without fracture/fracture dislocation/traumatic disc herniation/instability, there is requirement of high-quality prospective randomized controlled trials to resolve controversy regarding early surgery versus conservative management and delayed surgery if recovery plateaus or if there is a neurological deterioration. Until such time decision on surgery and its timing should be left to the judgment of physician, deliberating on pros and cons relevant to the particular patient and involving the well-informed patient and relatives in decision making. These slides can be retrieved under Electronic Supplementary Material.

Shunt migration in ventriculoperitoneal shunting: A comprehensive review of literature.

Ventriculoperitoneal shunts are one of the commonest surgical procedures performed in neurosurgery. Complications of this procedure include mechanical complications and nonmechanical ones. The most distressing mechanical complication is shunt migration, and often when this occurs, it becomes difficult to work out the mechanism and the management protocol. This review is designed to answer questions regarding management details of this unfortunate complication, and also seeks to identify the causes.

ISNO consensus guidelines for practical adaptation of the WHO 2016 classification of adult diffuse gliomas.

INTRODUCTION: Recent advances in the molecular biology of adult diffuse gliomas have brought about a paradigm shift in their diagnostic criteria, as witnessed in the World Health Organization (WHO) 2016 guidelines for central nervous system tumors. It is now mandatory to perform several molecular tests to reach a definitive integrated diagnosis in most of the cases. This comes with additional cost and higher turnaround time, which is not always affordable in developing countries like India. In addition, the non-uniform distribution of advanced research and diagnostic testing centers adds to the difficulty. METHODS: The Indian Society of Neuro-oncology (ISNO) multidisciplinary expert panel consisting of neuropathologists, neurosurgeons, and radiation/medical oncologists convened to prepare the national consensus guidelines for approach to diagnosis of adult diffuse gliomas. RESULTS: Algorithms for arriving at an integrated diagnosis of adult diffuse gliomas predominantly using immunohistochemistry and with minimum possible additional molecular testing were agreed upon, thus addressing the problems of cost, accessibility, and turnaround time. Mandatory and optional tests were proposed for each case scenario. CONCLUSION: This document represents the consensus of the various neuro-oncology disciplines involved in diagnosis and management of patients with adult diffuse gliomas. The article reflects a practical adaptation of the WHO recommendations to suit a resource constrained setup.

The spectrum of tuberculosis of the spine in pediatric age group: a review.

INTRODUCTION: Pediatric spinal tuberculosis is characterized by rapid bone destruction and carries the risk of rapid onset neurological deficits and severe deformity of the spine. Behavior of spinal deformity over time is affected by growth of spine. Owing to this dynamic behavior of pediatric spinal tuberculosis both in active phase and in healed phase, it presents with challenges which are quite different from adults with caries spine. A clinician must have high index of suspicion for accurate and early diagnosis of spinal tuberculosis in the pediatric population and should also have a thorough knowledge of differences in natural history between adult and pediatric spinal tuberculosis. DISCUSSION: This is based on the senior author's experience of dealing with tuberculosis of the spine in children over the last two decades. Recent advances in field of rapid diagnosis of tuberculosis based on nuclear material-related diagnostic tests have further improved the management of tuberculosis. At the same time, the basic treatment principles remain the same. However, the threshold for surgical vs conservative treatment have subtle differences when compared to adult population. The importance of long-term follow-up after treatment must be appreciated. CONCLUSION: Tuberculosis in the spine in children needs early attention. Prompting to diagnostic and medical therapy measures can avoid neurological sequellae and delayed deformity.

Indian Society of Neuro-Oncology consensus guidelines for the contemporary management of medulloblastoma.

INTRODUCTION: The high success rate in the management medulloblastoma achieved in the western world is not exactly mirrored in developing countries including India. Socio-demographic differences, health-care disparity, and lack in uniformity of care with resultant widespread variations in the clinical practice are some of the reasons that may partly explain this difference in outcomes. Patients with medulloblastoma require a multi-disciplinary team approach involving but not limited to neuro-radiology, neurosurgery; neuropathology, molecular biology, radiation oncology, pediatric medical oncology and rehabilitative services for optimizing outcomes. METHODS: The Indian Society of Neuro-Oncology (ISNO) constituted an expert multi-disciplinary panel with adequate representation from all stakeholders to prepare national consensus guidelines for the contemporary management of medulloblastoma. RESULTS: Minimum desirable, as well as preferable though optional recommendations (as appropriate), were developed and adopted for the pre-surgical work-up including neuroimaging; neurosurgical management including surgical principles, techniques, and complications; neuropathology reporting and molecular testing; contemporary risk-stratification in the molecular era; appropriate adjuvant therapy (radiotherapy and chemotherapy); and follow-up schedule in medulloblastoma. CONCLUSIONS: The current document represents a broad consensus reached amongst various stakeholders within the neuro-oncology community involved in the contemporary curative-intent management of children with medulloblastoma. It provides both general as well as specific guidelines and recommendations to be adopted by physicians and health care providers across India to achieve uniformity of care, improve disease-related outcomes, and compare results between institutions within the country.

A comprehensive and scalable database search system for metaproteomics.

BACKGROUND: Mass spectrometry-based shotgun proteomics experiments rely on accurate matching of experimental spectra against a database of protein sequences. Existing computational analysis methods are limited in the size of their sequence databases, which severely restricts the proteomic sequencing depth and functional analysis of highly complex samples. The growing amount of public high-throughput sequencing data will only exacerbate this problem. We designed a broadly applicable metaproteomic analysis method (ComPIL) that addresses protein database size limitations. RESULTS: Our approach to overcome this significant limitation in metaproteomics was to design a scalable set of sequence databases assembled for optimal library querying speeds. ComPIL was integrated with a modified version of the search engine ProLuCID (termed "Blazmass") to permit rapid matching of experimental spectra. Proof-of-principle analysis of human HEK293 lysate with a ComPIL database derived from high-quality genomic libraries was able to detect nearly all of the same peptides as a search with a human database (~500x fewer peptides in the database), with a small reduction in sensitivity. We were also able to detect proteins from the adenovirus used to immortalize these cells. We applied our method to a set of healthy human gut microbiome proteomic samples and showed a substantial increase in the number of identified peptides and proteins compared to previous metaproteomic analyses, while retaining a high degree of protein identification accuracy and allowing for a more in-depth characterization of the functional landscape of the samples. CONCLUSIONS: The combination of ComPIL with Blazmass allows proteomic searches to be performed with database sizes much larger than previously possible. These large database searches can be applied to complex meta-samples with unknown composition or proteomic samples where unexpected proteins may be identified. The protein database, proteomic search engine, and the proteomic data files for the 5 microbiome samples characterized and discussed herein are open source and available for use and additional analysis.

Exosome-mediated delivery of the intrinsic C-terminus domain of PTEN protects it from proteasomal degradation and ablates tumorigenesis.

PTEN mutation is a frequent feature across a plethora of human cancers, the hot-spot being its C-terminus (PTEN-CT) regulatory domain resulting in a much diminished protein expression. In this study, the presence of C-terminus mutations was confirmed through sequencing of different human tumor samples. The kinase CKII-mediated phosphorylation of PTEN at these sites makes it a loopy structure competing with the E3 ligases for binding to its lipid anchoring C2 domain. Accordingly, it was found that PTEN-CT expressing stable cell lines could inhibit tumorigenesis in syngenic breast tumor models. Therefore, we designed a novel exosome-mediated delivery of the intrinsic PTEN domain, PTEN-CT into different cancer cells and observed reduced proliferation, migration, and colony forming ability. The delivery of exosome containing PTEN-CT to breast tumor mice model was found to result in significant regression in tumor size with the tumor sections showing increased apoptosis. Here, we also report for the first time an active PTEN when its C2 domain is bound by PTEN-CT, probably rendering its anti-tumorigenic activities through the protein phosphatase activity. Therefore, therapeutic interventions that focus on PTEN E3 ligase inhibition through exosome-mediated PTEN-CT delivery can be a probable route in treating cancers with low PTEN expression.

Infant brain tumours: a tale of two cities.

INTRODUCTION: Infantile brain tumours (age < 1 year) are increasingly being diagnosed due to advances in prenatal and perinatal diagnostic imaging. We present here our retrospective study of 64 infant brain tumours that brings to the fore the epidemiology, clinical presentation, pathology and outcome of this unique subset of paediatric brain tumours presenting to two tertiary referral centres in Kolkata in India and Lille in France between the years 1999 and 2014. METHODS: Data was retrospectively collected from Kolkata (n = 30) and Lille (n = 34) for patients presenting with infant brain tumours and analysed for factors such as age at presentation, clinical features, gender, location of tumour, pathology, management and outcome. Follow-up was available for all patients. RESULTS: Mean age at presentation was 6.8 months at Kolkata and 6.3 months at Lille. More than two-thirds of tumours in both the groups were supratentorial and presented with signs of raised intracranial pressure. There was also a similar proportion of tumours presenting as congenital tumours. At Kolkata, germ cell tumours (n = 7) were the most common while low-grade gliomas (n = 11) formed the largest group at Lille. Kolkata had a higher incidence of high-grade gliomas (n = 5) and PNETs (n = 4) while ATRT (n = 3) and choroid plexus carcinoma (n = 4) were more common at Lille. Surgery was the mainstay of treatment at both centres. CONCLUSION: Brain tumours in infants presenting to tertiary centres in Europe and India are challenging to manage and usually have dismal prognosis. These tumours differ markedly in the pathology and, therefore, overall outcome. Surgery forms mainstay of treatment. Radiotherapy is best avoided in this age group.

Intracranial germ cell tumors: a multi-institutional experience from three tertiary care centers in India.

OBJECTIVE: Central nervous system germ cell tumors (CNS GCTs) are relatively rare neoplasms. Incidence of CNS GCTs in Western literature is low (0.3-0.6 %) as compared to East Asia (3-4 %). No large study is available on CNS GCTs from India. METHODS: Intracranial GCT cases were retrieved from databases of three tertiary care institutes in India; clinicopathological data was reviewed. RESULTS: Ninety-five intracranial GCT cases were identified, accounting for 0.43 % of CNS tumors. Median age was 12 years (range, birth to 48 years); male preponderance was noted (66 %). Most patients (86.3 %) were aged <18 years. Pineal location was most common (45 %) and was associated with male gender and age >14 years. Germinoma was the commonest histopathological type (63.2 %), followed by teratoma (20 %). Suprasellar location was associated with germinoma histology. Follow-up was available for 71 patients (median, 15 months). Of these, 48 received adjuvant chemotherapy and/or radiotherapy. At the last follow-up, 44 patients showed no evidence of disease. Age >10 years, male gender, pineal location, and germinoma histology were associated with favorable outcome. CONCLUSION: This is the first multicentric study from India establishing that incidence of CNS GCT in India is similar to that in the West and differs from that in East Asian countries. However, similar to both, germinoma is the commonest histological type, and pineal location is most frequent. Studies on molecular alterations based on ethnicity and geographical location are necessary to provide clarity on differences in incidence. Attention needs to be focused on decreasing treatment heterogeneity and minimizing treatment-related morbidity and mortality, improving the cure rate of these highly treatable tumors.