Diagnostic Utility of Next-Generation Sequencing in Circulating Free DNA and a Comparison With Matched Tissue in Gallbladder Carcinoma.

Mutation detection for therapy monitoring in cell-free DNA (cfDNA) is used clinically for some malignancies. Gallbladder carcinoma (GBC) presents a diagnostic challenge and has limited late-stage treatment options. To our knowledge, this novel study examines, for the first time, genomic alterations in cfDNA from GBC to assess diagnostic accuracy and therapeutic options. The concordance of somatic genomic changes in cfDNA and DNA from paired tumor tissue was analyzed. Paired serum and tissue samples from 40 histologically proven GBC, 20 cholecystitis, and 4 normal (noninflamed gallbladder) controls were included. Targeted next-generation sequencing with a 22-gene panel (Colon and Lung Cancer Research Panel v2, Thermo Scientific) in cfDNA and tumor tissue with high depth and uniform coverage on ION Personal Genome Machine (ION, PGM) was performed. A spectrum of 223 mutations in cfDNA and 225 mutations in formalin-fixed paraffin-embedded tissue DNA were identified in 22 genes. Mutations ranged from 1 to 17 per case. In cfDNA frequent alterations were in TP53 (85.0%), EGFR (52.5%), MET (35%) CTNNB1, SMAD4, BRAF (32.5%), PTEN (30%), FGFR3 and PIK3CA (27.5%), NOTCH1 (25.0%), and FBXW7 and ERBB4 (22.5%). At least one clinically actionable mutation was identified in all cfDNA samples. Paired samples shared 149 of 225 genetic abnormalities (66.2%). Individual gene mutation concordance ranged from 44.44% to 82.0% and was highest for EGFR (82.0%), BRAF and NOTCH1 (80.0%), TP53 (73.08%), MET (72.22%), and ERBB4 (71.42%) with a significant level of correlation (Spearman r = 0.91, P ≤ .0001). The sensitivity and specificity of the TP53 gene at the gene level was the highest (94.44% and 100.0%, respectively). Overall survival was higher for ERBB4 and ERBB2 mutant tumors. The adenocarcinoma subtype revealed specific genetic changes in ERBB4, SMAD4, ERBB2, PTEN, KRAS, and NRAS. NGS-based cfDNA mutation profiling can be used to diagnose GBC before surgery to guide treatment decisions. Targeted therapy identified in GBC included SMAD4, ERBB2, ERBB4, EGFR, KRAS, BRAF, PIK3CA, MET, and NRAS.

Biodegradation of polyethylene terephthalate (PET) by Brucella intermedia IITR130 and its proposed metabolic pathway.

Accumulation of polyethylene terephthalate (PET) polyester in ecosystems across the globe is a major pollution of concern. Microbial degradation recently generated novel insights into the biodegradation of varieties of plastics. In this study, a PET degrading bacterium Brucella intermedia IITR130 was isolated from a contaminated lake ecosystem at Pallikaranai, Chennai, India. Incubation of the bacterium along with the PET sheet (0.1 mm thickness) for 60 days resulted in 26.06% degradation, indicating a half-life of 137.8 days. Considerable changes in the surface morphology of the PET sheet were found as holes, pits, and cracks on incubation with strain IITR130, as revealed by scanning electron microscopy (SEM). After bacterial treatment of PET, the formation of new functional groups, most notably in the area of 3326 cm-1 suggestive of O-H stretch, leading to carboxylic acid and alcohol as products were suggested by fourier transform infrared (FTIR) analysis. Monomethyl terephthalate (MMT) and terephthalic acid (TPA) were identified by gas chromatography-mass spectrometry (GC-MS) analysis as PET degradation metabolites. Tributyrin clearance assay confirmed the presence of a lipase/esterase enzyme in the strain IITR130. In this study, a degradation pathway for PET by an isolated and identified bacterium Brucella intermedia IITR130 was characterized in detail.

Targeting Artemisinin-Resistant Malaria by Repurposing the Anti-Hepatitis C Virus Drug Alisporivir.

The emergence of Plasmodium falciparum resistance raises an urgent need to find new antimalarial drugs. Here, we report the rational repurposing of the anti-hepatitis C virus drug, alisporivir, a nonimmunosuppressive analog of cyclosporin A, against artemisinin-resistant strains of P. falciparum. In silico docking studies and molecular dynamic simulation predicted strong interaction of alisporivir with PfCyclophilin 19B, confirmed through biophysical assays with a Kd value of 354.3 nM. Alisporivir showed potent antimalarial activity against chloroquine-resistant (PfRKL-9 with resistance index [Ri] 2.14 ± 0.23) and artemisinin-resistant (PfKelch13R539T with Ri 1.15 ± 0.04) parasites. The Ri is defined as the ratio between the IC50 values of the resistant line to that of the sensitive line. To further investigate the mechanism involved, we analyzed the expression level of PfCyclophilin 19B in artemisinin-resistant P. falciparum (PfKelch13R539T). Semiquantitative real-time transcript, Western blot, and immunofluorescence analyses confirmed the overexpression of PfCyclophilin 19B in PfKelch13R539T. A 50% inhibitory concentration in the nanomolar range, together with the targeting of PfCyclophilin 19B, suggests that alisporivir can be used in combination with artemisinin. Since artemisinin resistance slows the clearance of ring-stage parasites, we performed a ring survival assay on artemisinin-resistant strain PfKelch13R539T and found significant decrease in parasite survival with alisporivir. Alisporivir was found to act synergistically with dihydroartemisinin and increase its efficacy. Furthermore, alisporivir exhibited antimalarial activity in vivo. Altogether, with the rational target-based Repurposing of alisporivir against malaria, our results support the hypothesis that targeting resistance mechanisms is a viable approach toward dealing with drug-resistant parasite.

A Review on Potential Footprints of Ferulic Acid for Treatment of Neurological Disorders.

Ferulic acid is being screened in preclinical settings to combat various neurological disorders. It is a naturally occurring dietary flavonoid commonly found in grains, fruits, and vegetables such as rice, wheat, oats, tomatoes, sweet corn etc., which exhibits protective effects against a number of neurological diseases such as epilepsy, depression, ischemia-reperfusion injury, Alzheimer's disease, and Parkinson's disease. Ferulic acid prevents and treats different neurological diseases pertaining to its potent anti-oxidative and anti-inflammatory effects, beside modulating unique neuro-signaling pathways. It stays in the bloodstream for longer periods than other dietary polyphenols and antioxidants and easily crosses blood brain barrier. The use of novel drug delivery systems such as solid-lipid nanoparticles (SLNs) or its salt forms (sodium ferulate, ethyl ferulate, and isopentyl ferulate) further enhance its bioavailability and cerebral penetration. Based on reported studies, ferulic acid appears to be a promising molecule for treatment of neurological disorders; however, more preclinical (in vitro and in vivo) mechanism-based studies should be planned and conceived followed by its testing in clinical settings.

Panel of serum long non-coding RNAs as potential non-invasive biomarkers for gallbladder carcinoma.

Gallbladder carcinoma (GBC) is a common malignancy and is usually diagnosed in the late stages of the disease. The identification of new effective early diagnostic biomarkers could represent an effective approach in reducing mortality in GBC. Altered expression of long non-coding RNAs (lncRNAs) is believed to be associated with the emergence and development of GBC. Our study aims to identify the expression of a range of circulating lncRNAs, including HOTAIR, ANRIL, H19, CCAT1 and MEG3, in matched serum and tissues of GBC for diagnosis and its association with clinicopathological features. The case and control study included matched serum and tissues from 63 GBC, 19 cholecystitis (CC), and 46 normal controls (NC). RNA extraction and cDNA synthesis from serum and fresh tissue match were performed using commercially available kits. Relative expression was assessed using SYBR Green real-time quantitative polymerase chain reaction. Circulating lncRNA levels including HOTAIR, ANRIL and H19 were upregulated in serum samples, while MEG3 and CCAT1 were downregulated in GBC compared to controls. The trend towards upregulation and downregulation was comparable in the tissue. HOTAIR and MEG3 levels were significantly different between serum CC and early-stage GBC (p = 0.0373, 0.0020), while H19 was significantly upregulated comparing early-stage GBC to advanced-stage GBC (p = 0.018). The expression of ANRIL was significant with M stage (p = 0.0488), H19 with stage (p = 0.009), M stage (p=<0.0001) & stage (0.009) and CCAT1 with M stage (0.044). When distinguishing GBC and NC, AUC for HOTAIR was 0.75, ANRIL 0.78, H19 0.74, CCAT1 0.80 and 0.96 for MEG3. The combination sensitivity for lncRNAs ranged from 84.13% (CI: 72.74-92.12%) to 100.0% (CI: 94.31-100.0%). Significant diagnostic value in discriminating pathologic stage was observed for ANRIL and MEG3 (p = 0.022, p = 0.0005). LncRNA show a significant change in expression in GBC and in discrimination of early stage from late-stage disease. The detection of 2 lncRNAs in panels, in coordination with radiology, could represent a potential serum-based biomarker for early-stage GBC diagnosis.

Covid-19-Associated Mucormycosis: Histopathology of the Deadly Fungal Infection.

Introduction Many patients suffered from rhino-orbital-cerebral mucormycosis during the coronavirus disease 2019 (COVID-19) pandemic in India. Diabetes is a known risk factor of COVID-19 infection and mucormycosis. Objective The present study was done to describe the clinical spectrum and histopathological findings of mucormycosis in COVID-19 patients and their outcomes. Methods A cross-sectional study was done over a period of two and half months. The biopsy samples or scrapings from sinonasal or periorbital tissue of 38 patients were analyzed. Hematoxylin & Eosin (H&E stain) slides were evaluated along with Grocott-Gomori methenamine-silver and Periodic acid-Schiff stains to highlight the fungal elements. Results The male to female ratio was 2.5:1, and the mean age of the subjects was 53 years old. A total of 68.4% ( n = 26/38) of the patients had diabetes as a comorbidity, 84.2% ( n = 32/38) had a history of steroid intake, and 55.3% ( n = 21/38) were given supplemental oxygen during their treatment. The common presentations were nasal blockage, discharge, eye pain, headache, and altered mentation. The sites of biopsy were: nasal cavity 76.3% ( n = 29/38), periorbital fat/orbit 21.1% ( n = 8/38), maxillary sinus 15.8% ( n = 6/38) and ethmoid sinus 13.2% ( n = 5/38). In 76.3% ( n = 29/38) cases, broad, irregular, nonseptate, and right-angle branching hyphae were seen on H&E-stained tissue sections. Conclusion COVID-19 led to various complications in individuals affected by it. Mucormycosis was one such lethal complication. An early diagnosis and prompt treatment is crucial to control the progression of the disease and improve outcomes.

Identification of a De Novo Peptide against Palmitoyl Acyltransferase 6 to Block Survivability and Infectivity of Leishmania donovani.

Palmitoylation is an essential post-translational modification in Leishmania donovani, catalyzed by enzymes called palmitoyl acyl transferases (PATs) and has an essential role in virulence. Due to the toxicity and promiscuity of known PAT inhibitors, identification of new molecules is needed. Herein, we identified a specific novel de novo peptide inhibitor, PS1, against the PAT6 Leishmania donovani palmitoyl acyl transferase (LdPAT6). To demonstrate specific inhibition of LdPAT6 by PS1, we employed a bacterial orthologue system and metabolic labeling-coupled click chemistry where both LdPAT6 and PS1 were coexpressed and displayed palmitoylation suppression. Furthermore, strong binding of the LdPAT6-DHHC domain with PS1 was observed through analysis using microscale thermophoresis, ELISA, and dot blot assay. PS1 specific to LdPAT6 showed significant growth inhibition in promastigotes and amastigotes by expressing low cytokines levels and invasion. This study reveals discovery of a novel de novo peptide against LdPAT6-DHHC which has potential to block survivability and infectivity of L. donovani.

A rare case of left atrial myxofibromyosarcoma.

Myxofibrosarcoma is an aggressive soft tissue sarcoma, previously known as myxoid variant of malignant fibrous histiocytoma. Primary cardiac myxofibrosarcomas are the rarest forms of cardiac malignant tumors that often remain asymptomatic until metastasis occurs. In this case report, we describe a rare left atrial cardiac myxofibrosarcoma in a patient with recurrent renal cell carcinoma. We discuss the multimodality imaging approach to diagnose and evaluate cardiac masses as well as imaging characteristics to differentiate cardiac masses from thrombus.

Next Generation Sequencing in Vitellointestinal Duct Adenoma: Existence of Adenoma-Carcinoma Sequence or too Early to Predict?

Patent vitello-intestinal duct with adenoma is rare presentation. We report a case of a 1-month-old boy presenting with intermittent passage of stool and blood from the umbilicus since birth. On local examination polypoidal mass measuring 1×1 cm was seen protruding from umbilicus with faecal discharge. Ultrasound was performed which revealed a tubular hyperechoic structure, extending from umbilicus to part of small intestine measuring 30 ×30 mm and clinical diagnosis of patent vitello-intestinal duct was given, exploratory laparotomy, excision with umbilicoplasty done, and send for histopathological examination. On histopathological examination, patent vitello-intestinal duct adenoma was rendered and next generation sequencing (NGS) was performed revealing somatic mutation of KRAS (NM_033360.4; c.38G>A; p.Gly12Asp). To our knowledge, this is the first report of the adenoma in patent vitello-intestinal duct with NGS analysis. This case emphasizes the importance of thorough microscopic examination of resected patent vitello-intestinal duct and mutational analysis of the early lesions.

Anorectal balloon cell melanoma: a rare variant.

Balloon cell melanoma is a rare presentation of malignant melanoma, usually on the skin, with less than 100 cases reported. Mucosal BCM is even rarer, with only one case of anorectal BCM reported in English literature. The diagnosis is based on the histopathologic findings of a tumor composed of large, foamy melanocytes, with or without pigmentation, and confirmed by immunohistochemical studies showing expression for melanocytic markers. The foam cell appearance of the tumor cells and the lack of melanin pigment lead to a diagnostic dilemma, mostly when presented at an unusual location. Herein, we report a case of balloon cell melanoma at the anorectal junction in a 73-year-old male patient complaining of constipation and bleeding per rectum. Surgical resection was performed with no evidence of recurrence after three years of close follow-up. We believe this case will raise awareness among the medical community to consider this tumor a differential diagnosis in rectal masses.

Circulating microRNAs in gallbladder cancer: Is serum assay of diagnostic value?

The microRNAs (miRNAs) in circulation could serve as biomarkers for cancer detection. Gallbladder carcinoma (GBC) is mostly asymptomatic; therefore, using microRNAs (miRNAs) as an early diagnostic biomarker could be a valuable tool. We aimed to identify the tumor-associated miR-1, miR130, miR-146, miR-182, and miR-21expression in serum as a biomarker for early detection of GBC and identify their possible diagnostic role. The study group comprised of paired serum and tissue samples from 34 GBC, 19 cholecystitis (CC), 21 normal controls (uninflamed gall bladder), and additional 29 serum-only samples of GBC. Total RNA was isolated using a commercially available RNA isolation kit (Applied Biosystem, USA) and reverse transcribed using Advanced Taqman MicroRNA reverse transcription kit. The relative expression of miRNAs was analyzed using Quantitative real-time polymerase chain reaction. The diagnostic potential of these miRNAs was assessed by ROC analysis. In paired samples, the trend towards up and down regulation for miR-182, miR-21, miR-1, miR-130, and miR-146 was similar in both tissue and sera of GBC. The expression pattern of serum miR-1, miR130, and miR-146 gradually decreased from normal control (NC) to CC to GBC, while miR-21 and miR-182 gradually increased from NC to CC to GBC. The miR-1, miR-121, miR-182, and miR-146 significantly differed between CC vs. early stage and early stage vs. NC. Among these miRNAs, the sensitivity of miR-1 (85.71 %) was the highest, and the specificity of miR-21 was the highest (92.73 %). The combined sensitivity for miRNAs ranged from 73.13 % (CI: 60.90-83.24 %) to 98.63 % (CI: 89.0-99.61 %); however, the specificity was lower. In stage I&II vs. III&IV discrimination, the diagnostic sensitivity of miR-1 was highest (89.36 %, CI: 76.90-96.45). The two miRNAs, in combination, increase the diagnostic sensitivity. Circulating serum miRNAs may provide a new approach for clinical application. Panels of specific circulating miRNA, which require further validation, could be potential non-invasive diagnostic biomarkers for GBC in combination with abnormal radio diagnostic scans.

Concomitant Non-V600E BRAF and KRAS Mutations in Colorectal Carcinoma by Next-Generation Sequencing: A Distinct Subtype.

The RAS-RAF-MEK-ERK signaling cascade is the most frequently affected signaling pathway in colorectal cancer. BRAFV600E mutations serve as a drug-treatable hotspot and KRAS mutations as a predictor of susceptibility to anti-epidermal growth factor receptor therapy. Concomitant non-V600E BRAF and KRAS mutations may coexist and are rarely reported in the literature. We report a patient of colorectal carcinoma with inguinal lymph node metastases harboring mutations at the KRAS and BRAF non-V600E mutation codon detected by next-generation sequencing with an emphasis on clinical, pathological, and therapeutic implications of the mutation and review of the literature.

Defined Microenvironments Trigger In Vitro Gastrulation in Human Pluripotent Stem Cells.

Gastrulation is a stage in embryo development where three germ layers arise to dictate the human body plan. In vitro models of gastrulation have been demonstrated by treating pluripotent stem cells with soluble morphogens to trigger differentiation. However, in vivo gastrulation is a multistage process coordinated through feedback between soluble gradients and biophysical forces, with the multipotent epiblast transforming to the primitive streak followed by germ layer segregation. Here, the authors show how constraining pluripotent stem cells to hydrogel islands triggers morphogenesis that mirrors the stages preceding in vivo gastrulation, without the need for exogenous supplements. Within hours of initial seeding, cells display a contractile phenotype at the boundary, which leads to enhanced proliferation, yes-associated protein (YAP) translocation, epithelial to mesenchymal transition, and emergence of SRY-box transcription factor 17 (SOX17)+ T/BRACHYURY+ cells. Molecular profiling and pathway analysis reveals a role for mechanotransduction-coupled wingless-type (WNT) signaling in orchestrating differentiation, which bears similarities to processes observed in whole organism models of development. After two days, the colonies form multilayered aggregates, which can be removed for further growth and differentiation. This approach demonstrates how materials alone can initiate gastrulation, thereby providing in vitro models of development and a tool to support organoid bioengineering efforts.

Anti-inflammatory and neuroprotective effects of an orally active apocynin derivative in pre-clinical models of Parkinson's disease.

BACKGROUND: Parkinson's disease (PD) is a devastating neurodegenerative disorder characterized by progressive motor debilitation, which affects several million people worldwide. Recent evidence suggests that glial cell activation and its inflammatory response may contribute to the progressive degeneration of dopaminergic neurons in PD. Currently, there are no neuroprotective agents available that can effectively slow the disease progression. Herein, we evaluated the anti-inflammatory and antioxidant efficacy of diapocynin, an oxidative metabolite of the naturally occurring agent apocynin, in a pre-clinical 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. METHODS: Both pre-treatment and post-treatment of diapocynin were tested in the MPTP mouse model of PD. Diapocynin was administered via oral gavage to MPTP-treated mice. Following the treatment, behavioral, neurochemical and immunohistological studies were performed. Neuroinflammatory markers, such as ionized calcium binding adaptor molecule 1 (Iba-1), glial fibrillary acidic protein (GFAP), gp91phox and inducible nitric oxide synthase (iNOS), were measured in the nigrostriatal system. Nigral tyrosine hydroxylase (TH)-positive neurons as well as oxidative markers 3-nitrotyrosine (3-NT), 4-hydroxynonenal (4-HNE) and striatal dopamine levels were quantified for assessment of the neuroprotective efficacy of diapocynin. RESULTS: Oral administration of diapocynin significantly attenuated MPTP-induced microglial and astroglial cell activation in the substantia nigra (SN). MPTP-induced expression of gp91phox and iNOS activation in the glial cells of SN was also completely blocked by diapocynin. Notably, diapocynin markedly inhibited MPTP-induced oxidative markers including 3-NT and 4-HNE levels in the SN. Treatment with diapocynin also significantly improved locomotor activity, restored dopamine and its metabolites, and protected dopaminergic neurons and their nerve terminals in this pre-clinical model of PD. Importantly, diapocynin administered 3 days after initiation of the disease restored the neurochemical deficits. Diapocynin also halted the disease progression in a chronic mouse model of PD. CONCLUSIONS: Collectively, these results demonstrate that diapocynin exhibits profound neuroprotective effects in a pre-clinical animal model of PD by attenuating oxidative damage and neuroinflammatory responses. These findings may have important translational implications for treating PD patients.

Attenuation of Aß-induced neurotoxicity by thymoquinone via inhibition of mitochondrial dysfunction and oxidative stress.

Beta-amyloid (Aß) peptides are considered to play a major role in the pathogenesis of Alzheimer's disease (AD) and compounds that can prevent pathways of Aß-induced neurotoxicity may be potential therapeutic agents for treatment of AD. This study examined the hypothesis that thymoquinone (TQ) would reduce oxidative stress and mitochondrial dysfunction in differentiated pheochromocytoma (PC 12) cells exposed to Aß fragment 25-35 (Aß(25-35)). To test this hypothesis, Aß was used to induce an in vitro model of AD in differentiated PC 12 cell line of rat. After 24 h of exposure with Aß(25-35), a significant reduction in cell viability and mitochondrial membrane potential (MMP) was observed. In addition, a significant elevation in the TBARS content and nitric oxide (NO) and activity of acetylcholine esterase (AChE) was observed which was restored significantly by TQ pretreatment. Furthermore, TQ also ameliorated glutathione and its dependent enzymes (glutathione peroxidase, glutathione reductase) which were depleted by Aß(25-35) in PC 12 cells. These results were supported by the immunocytochemical finding that has shown protection of cells by TQ from noxious effects of Aß(25-35). These results indicate that TQ holds potential for neuroprotection and may be a promising approach for the treatment of neurodegenerative disorders including AD.

Liquid biopsy in CNS tumors: Current status & future perspectives.

Precise classification of central nervous system (CNS) malignancies is vital for the treatment and prognostication. Identification of noninvasive markers can be of importance to guide treatment decisions and in monitoring treatment response. CNS tumors are classified based on morphology with an essential complement of molecular changes, including mutations, amplifications, and methylation. Neuroimaging is the mainstay for initial diagnosis and monitoring tumor response with obvious limitations of imprecise tumor typing and no information on diagnostic, predictive and prognostic markers. Liquid biopsy has evolved as a diagnostic tool in body fluids and is being investigated as a surrogate for tissue biopsy in managing primary and metastatic brain tumors. Liquid biopsy refers to analyzing biological fluids such as peripheral blood, urine, pleural effusion, ascites, and cerebrospinal fluid (CSF); however, peripheral blood remains the primary source of fluid biopsy. The analytes include cell-free DNA (cfDNA) circulating tumor cells (CTCs), circulating micro RNAs (miRNAs), circulating proteins and extracellular vesicles (EVs). Analysis of these components is actively used for early cancer detection, auxiliary staging, prognosis assessment, detection of minimal residual disease (MRD), and monitoring drug resistance in various solid tumors. In recent years, liquid biopsy has been studied in CNS tumors, and analysis of CTCs and cfDNA have become relevant research topics. In the current review, we have explained the clinical potential of liquid biopsy in CNS tumors to assist in diagnosing and predicting prognosis and response to treatment.

The role of pharmaceutical industry in building resilient health system.

Objectives: This study explores the interrelationship among the current sustainability agenda of the pharmaceutical industry, based on the United Nation sustainable development goals (SDGs), the elements of the Joint External Evaluation (JEE) tool, and the triad components of the One Health approach. Methods: A cross-walk exercise was conducted to identify commonalities among SDGs, JEE assessment tool, and One Health approach. An in-depth study of 10 global pharmaceutical firms' corporate sustainability reports and COVID-19 response plan for 2019-2020 was also conducted. Results: The result of the exercise showed the existence of a direct and indirect relationship among the SDGs, elements of JEE assessment tool, and One Health approach. For example, both no poverty (SDG 1) and zero hunger (SDG 2) are linked with food safety targets under the JEE and with human and animal health under the One Health approach. Conclusion: This study adds a new dimension emphasizing the possibility of tailoring the pharmaceutical industry's activities under the sustainability agenda to strengthen global health security while remaining consistent with the One Health approach.

Cytomorphology of pituitary carcinoma metastatic to liver diagnosed by fine-needle aspiration: A rare case report and review of literature.

Pituitary carcinomas (PCs) are rare entities constituting about 0.1-0.2% of all pituitary neoplasms. They are diagnosed by the presence of craniospinal or systemic metastasis in pituitary adenomas (PAs). The distant metastatic sites include liver, followed by bone, lung, and lymph nodes. The diagnosis of PC is rarely made on fine-needle aspiration cytology (FNAC) with only six cases reported till date; hence, the cytologic features are not well defined. Herein, we report a case of PA having high Ki-67 proliferation index and p53 expression, presenting with liver lesion 6 weeks post-surgery and diagnosed on FNA. Detailed cytomorphologic features are defined and compared. We emphasize that FNAC, along with clinic-radiologic correlation, is a cost-effective, safe, and diagnostically accurate method of diagnosing PC metastases.

Genomic profiling of gallbladder carcinoma: Targetable mutations and pathways involved.

Gallbladder cancer (GBC) carries a poor prognosis and frequently present in late stages of disease. Identification of targetable molecular changes for selecting appropriate treatment and identifying prognostic and therapeutic subsets is required. Next-generation sequencing (NGS) using a frequently mutated for GBC may provide a reference for clinical management. The primary aim of the current study was to analyze the frequency of individual genetic alterations in GBC and to correlate with clinicopathological characteristics. A retrospective study was conducted using 22 gene panel examined NGS based approach for the detection of actionable genomic mutations in 37 GBC patients. The genetic and clinicopathological characteristics were analyzed. The number of mutations in cases was ranged from 1 to 15. A total of 171 mutations were identified in FFPE tissue DNA. The most common alterations were TP53 (90.90%), SMAD4 (60.60%), NOTCH1(45.45)& ERBB2 (45.45%), PIK3CA (33.33%) and MET (30.30)& PTEN (30.30%). Among the 22 gene panel, the TP53 gene was associated with histopathological differentiation (p = 0.0001), ERBB4 & ALK mutation was associated with necrosis (p = 0.012, 0.027), EGFR mutation was associated with mucin status (p = 0.023) and ERBB2 gene mutation was associated with T stage (p = 0.036). The study provides an overview of the genetic alterations in GBC patients.Targetable mutations are present in 89.91% cases of GBC which include SMAD4, NOTCH1, ERBB2&4, PIK3CA, MET, PTEN, EGFR, KRAS and BRAF. Metastatic disease was associated with high frequency of CTNNB1, KRAS and NRAS gene mutations.

Biomaterials directed activation of a cryostable therapeutic secretome in induced pluripotent stem cell derived mesenchymal stromal cells.

Mesenchymal stem cell therapy has suffered from wide variability in clinical efficacy, largely due to heterogeneous starting cell populations and large-scale cell death during and after implantation. Optimizing the manufacturing process has led to reproducible cell populations that can be cryopreserved for clinical applications. Nevertheless, ensuring a reproducible cell state that persists after cryopreservation remains a significant challenge, and is necessary to ensure reproducible clinical outcomes. Here we demonstrate how matrix-conjugated hydrogel cell culture materials can normalize a population of induced pluripotent stem cell derived mesenchymal stem cells (iPSC-MSCs) to display a defined secretory profile that promotes enhanced neovascularization in vitro and in vivo. Using a protein-conjugated biomaterials screen we identified two conditions-1 kPa collagen and 10 kPa fibronectin coated polyacrylamide gels-that promote reproducible secretion of pro-angiogenic and immunomodulatory cytokines from iPSC-MSCs that enhance tubulogenesis of endothelial cells in Geltrex and neovascularization in chick chorioallantoic membranes. Using defined culture substrates alone, we demonstrate maintenance of secretory activity after cryopreservation for the first time. This advance provides a simple and scalable approach for cell engineering and subsequent manufacturing, toward normalizing and priming a desired cell activity for clinical regenerative medicine.