A Study of Arginase Expression in Chronic Non-healing Wounds.

Arginase expression has been recently shown to increase in numerous disease states like neurodegeneration, inflammation, and malignancies. Although it has been found to be functionally important in various disease pathologies, little is known about its role in wound healing. Here, we look at the expression of arginase and its isoforms in chronic non-healing wounds and also study the expression of nitric oxide synthase (NOS) and oxidative stress enzymes in them. Wound tissues and blood samples were collected at the time of index presentation and follow-up from 61 chronic non-healing wound cases. The expression patterns of arginase isoenzymes, NOS, superoxide dismutases (SOD), lactic acid dehydrogenase (LDH), and catalase were examined by using enzyme-linked immunosorbent assay, immunohistochemistry, and western blot analysis at the transcript and protein level. We reported a significant decrease of serum arginase levels in chronic nonhealing wounds in the progress of wound healing. Interestingly, tissue arginase levels were found to be increased with improved wound condition at follow-up. Tissue NOS, LDH, and catalase activity were also found to be increased with the progress of healing, whereas SOD levels were downregulated. Our findings reported increased expression at the transcript level of arginase-I and arginase-II in chronic non-healing wounds for the first time. In conclusion, we observed decreased serum arginase levels in completely healed patients as compared to non-healed cases. Our study findings support the hypothesis that inhibition of the activity of arginase delays wound healing. Arginase and iNOS may also find their place in the future as possible biomarkers for wound healing.

Unravelling of the comparative Transcriptomic Profile of Gallbladder Cancer using mRNA sequencing.

BACKGROUND: Gallbladder cancer (GBC) represents a wide geographical diversity as well as heterogeneity in clinical and genomic landscape. There seems to be little progress in the development of diagnostic biomarkers, targeted therapies or individualized approaches to GBC management. In this study, we investigated the whole transcriptome profile of GBC patients using RNA sequencing and identified key genes and pathways associated with gallbladder cancer using bioinformatics. METHODOLOGY: A total of 10 cases of GBC were collected and sequenced. The raw reads of the gallbladder sample was compared with the gallbladder normal control (SRA Database ID: ERX288537: HPA RNA-seq normal tissues gallbladder). Using Gene ontology analysis the differentially expressed genes were categorized into the biological pathway, cellular component, and molecular function. Pathway enrichment analyses, protein-protein interaction, transcription factor and miRNA interaction that regulate the expression of hub genes were conducted using bioinformatics tool. RESULTS: A total of 954 differentially expressed mRNA transcripts were identified, including overexpression of REG4, TMEM238, S100A2, LYPD2, and KRT17, as well as underexpressed genes like CCKAR, IGSF10, CHRM2, CRISP3, and FGF19. Enrichment analysis showed the metabolic pathways to be the top five cancer pathways in gallbladder carcinogenesis besides PI3k-Akt signalling pathway, cAMP signalling pathway, miRNAs in cancer, and cell adhesion profile of GBC. CONCLUSIONS: CCKAR, CDKN2A and LRRK2 were found to be most involved genes in its progression and development through different regulatory pathways. Further, most of the genes were significantly involved in PI3k-Akt, Wnt and hedgehog signaling pathways which have a key role in gallbladder cancer development.

Study on Blood Serum Levels of Heavy and Trace Metals in Chronic Non-Healing Wounds.

Wound healing is a complex, highly regulated process that is important in sustaining the skin barrier function. The etiologic relation of specific metals is not adequately described for chronic non-healing wounds. The aim of this study was to estimate heavy and trace metals in chronic non-healing wound and their association with wound healing. The levels of zinc, selenium, copper, magnesium, chromium, cadmium, iron, and lead were estimated in serum of chronic non-healing wound patients using atomic absorption spectrophotometry. The tests were carried out in 50 patients with chronic non-healing wound and thirty healthy volunteers as control. The serum levels of elements namely zinc, selenium, copper, magnesium, and chromium were significantly reduced in chronic non-healing wounds (P < .001) as compared to control. Lead and cadmium levels had shown the significantly increasing trend in chronic non-healing wound cases (P < .001). The present study demonstrated a significant decrease in serum, levels of selenium, zinc, copper, magnesium, iron, and chromium levels in patients with chronic non-healing wound indicating an association between these elements and wound healing. To summarize the findings of our research, hence trace elements were decreasing in chronic non-healing wound patients suggesting their role in wound healing.

A systematic review of proteomic biomarkers in oral squamous cell cancer.

BACKGROUND: Head and neck squamous cell cancer (HNSCC) is the most common cancer associated with chewing tobacco, in the world. As this is divided in to sites and subsites, it does not make it to top 10 cancers. The most common subsite is the oral cancer. At the time of diagnosis, more than 50% of patients with oral squamous cell cancers (OSCC) had advanced disease, indicating the lack of availability of early detection and risk assessment biomarkers. The new protein biomarker development and discovery will aid in early diagnosis and treatment which lead to targeted treatment and ultimately a good prognosis. METHODS: This systematic review was performed as per PRISMA guidelines. All relevant studies assessing characteristics of oral cancer and proteomics were considered for analysis. Only human studies published in English were included, and abstracts, incomplete articles, and cell line or animal studies were excluded. RESULTS: A total of 308 articles were found, of which 112 were found to be relevant after exclusion. The present review focuses on techniques of cancer proteomics and discovery of biomarkers using these techniques. The signature of protein expression may be used to predict drug response and clinical course of disease and could be used to individualize therapy with such knowledge. CONCLUSIONS: Prospective use of these markers in the clinical setting will enable early detection, prediction of response to treatment, improvement in treatment selection, and early detection of tumor recurrence for disease monitoring. However, most of these markers for OSCC are yet to be validated.

mRNA Expression Analysis of E-Cadherin, VEGF, and MMPs in Gastric Cancer: a Pilot Study.

Gastric cancer (GC) is a serious fatal cancer on a global scale because of its presentation at advanced stage. The expressions of vascular endothelial growth factor (VEGF), E-cadherin, and matrix metalloproteinases (MMPs) in other cancers have been reported. However, its expression and underlying mechanisms are little known in gastric cancer in Indian context. In this study, we detected mRNA expression of VEGF, E-cadherin, and MMPs (MMP-1, MMP-2, and MMP-9) in 73 gastric cancer tissues and 27 normal controls by reverse-transcriptase polymerase chain reaction (RT-PCR). Receiver operator characteristics analysis was done for determining the diagnostic utility of VEGF, MMPs and E-cadherin with respect to the sensitivity and specificity. The association of VEGF, MMPs, and E-cadherin expression with the clinicopathological characteristics and the prognosis was subsequently analyzed. The mRNA expression results showed that E-cadherin was significantly downregulated in 47.9% of GC in comparison to control. There was no change in VEGF expression observed in 90.4% GC cases. MMP-1, MMP-2, and MMP-9 were overexpressed in 13.7%, 28.8%, and 11% of GC, respectively, with significant change in MMP-2 (p ≤ 0.0001) and MMP-9 (p = 0.027) in comparison to control. Our results strengthen the necessity of more studies to elucidate the prophetic role of these genes in the development of gastric cancer.

Analysis of SET and MYND Domain-Containing Protein 3 (SMYD3) Expression in Gallbladder Cancer: a Pilot Study.

The Suvar, Enhancer of zeste, and Trithorax (SET) and myeloid-Nervy-DEAF-1 (MYND) domain-containing protein 3 (SMYD3) is a histone lysine methyltransferase and has been recently unveiled to play significant roles in the progression of human cancer via regulating various key cancer-associated genes and pathways. The role of SMYD3 in gallbladder cancer (GBC) still needs to be studied. In the present study, we examined the SMYD3 gene expression at mRNA and protein level to look its impact on risk for developing gallbladder carcinogenesis. SMYD3 expression was evaluated by immunohistochemistry and reverse transcriptase PCR (RT-PCR) from 30 cases each of GBC and cholelithiasis patients. The expression was compared with different clinicopathological parameters. The SMYD3 expression was found to be significantly upregulated in GBC than cholelithiasis group (p < 0.05). The SMYD3 with increased expression level was observed in 73.3% of the GBC cases (p < 0.05). Moreover, mRNA SMYD3 expression was observed in 73.3% of GBC and 10% of control (p < 0.05). Our results indicated that the overexpression of SMYD3 plays an important role in the GBC progression, and SMYD3 may represent useful biomarker for gallbladder cancer patients.

Expression analysis of aryl hydrocarbon receptor repressor (AHRR) gene in gallbladder cancer.

BACKGROUND: The aryl hydrocarbon receptor repressor (AHRR), a member of the growing superfamily, is a basic helix-loop-helix/PerAHR nuclear translocator (ARNT)-Sim (bHLH-PAS) protein. AHRR has been proposed to function as a putative new tumor suppressor gene based on studies in multiple types of human cancers. This current study aims to investigate AHHR expression and its prognostic significance in gallbladder cancer. METHODS: The study includes 48 gallbladder cancer and 34 chronic cholecystitis cases as controls. The expression level of AHRR was analyzed by using semi-quantitative PCR and immunohistochemical staining. The results were correlated with different clinical parameters. RESULTS: We demonstrate that the expression of AHRR is significantly down-regulated in gallbladder cancer tissue samples as compared to that in chronic cholecystitis tissue samples by reverse transcriptase PCR (RT-PCR) (P = 0.017) and immunohistochemistry analysis (P = 0.002). Interestingly, our RT-PCR data revealed that AHRR mRNA expression is frequently down-regulated (45.8%; 22/48) in cases as compared to 14.7% (5/34) in controls. Similarly, immunohistochemical analysis data show significant down-regulation of AHRR expression in 77.1% (37/48) of gallbladder cancer cases than 44.1% (15/34) in controls (P < 0.017). Reduced mRNA and protein expression is significantly associated with advanced T-stage (P = 0.001), histological differentiation (P = 0.001), and tumors with nodal metastasis (P = 0.001). Decreased expression of AHRR is significantly associated with poor prognosis in gallbladder cancer patients. CONCLUSION: In conclusion, the present study suggests that low AHRR expression may be critical in gallbladder cancer development. Our data suggests that AHRR may act as a tumor suppressor gene and its expression profile may be useful as a diagnostic marker in gallbladder cancer.

Genetic mutational analysis of ß-catenin gene affecting GSK-3ß phosphorylation plays a role in gallbladder carcinogenesis: Results from a case control study.

This manuscript has reported different mutations of ß-catenin gene in gallbladder cancer patients which affect GSK-3ß phosphorylation site. PURPOSE: Gallbladder carcinoma (GBC) is a relatively rare and fatal cancer with poor prognosis. The molecular mechanism of gallbladder carcinogenesis is still not clear. Wnt signaling pathway is a highly conserved pathway that regulates proliferation, differentiation, migration, genetic stability, apoptosis, and stem cell renewal. ß-catenin plays major role in Wnt signaling and aberrations in ß-catenin has found to be involved in several cancers pathogenesis. This study was carried out to document the mutations of ß-catenin gene in gallbladder cancer and to evaluate its possible role in gallbladder carcinogenesis. METHODS: PCR-SSCP (Single Stranded Conformation Polymorphism) for ctnnb1 was performed in 50 patients each of gallbladder cancer, cholelithiasis and 50 healthy controls. Samples that showed variation in banding pattern were sequenced. RESULTS: Variation in banding pattern was observed in 9 (18%) samples of GBC, 4 (8%) of cholelithiasis and 2 (4%) of control. Sequencing analysis showed 9 novel mutations of ctnnb1 in exon 3 in 18% of gallbladder cancer (χ2 = 5.778; p < 0.05). Six point mutations, 1 deletion and 1 insertion mutation were found in 9 cases of gallbladder cancer. All point mutations were mis-sense mutation that affected highly conserved serine or threonine region that is important for GSK-3ß phosphorylation. CONCLUSION: Findings of the study suggests that high frequency of non synonymous mutations of ß-catenin gene (ctnnb1) occurs in patients with gallbladder cancer. As these mutations mainly effect GSK 3ß phosphorylation, it may be concluded that this might be an important step in gallbladder carcinogenesis. These ß-catenin mutations lead to Wnt pathway activation and appear to have a role in progression from inflammation to cancer in gallbladder.

Effect of neoadjuvant chemotherapy and its correlation with HPV status, EGFR, Her-2-neu, and GADD45 expression in oral squamous cell carcinoma.

BACKGROUND: Head and neck cancers are the commonest cancer in Southeast Asia. Despite being a surface cancer, it is associated with significant morbidity as despite early detection by the patients they often report for treatment late and hence are associated with poor prognosis. The role of neoadjuvant chemotherapy in head and neck cancer is still under evaluation; there is a large subgroup of population that does not respond to chemotherapy, and hence, most studies have failed to show any survival benefit. This study evaluated the role of neoadjuvant therapy with docetaxel and carboplatin in patients with oral cancer and correlated the response to human papilloma virus, EGFR1, EGFR2, and GADD45 expression. METHODS: A total of 24 locally advanced, non-metastatic oral cancer patients were included in the study. Tumor biopsies were taken prior to the start of neoadjuvant therapy for expression of EGFR, Her-2-Neu, and GADD45 by immunohistochemistry and for HPV by PCR. The response was evaluated using Response Evaluation Criteria in Solid Tumors (RECIST) criteria after three cycles of chemotherapy. Statistical analysis was performed using correlation and Kaplan-Meier analysis; the difference in survival was calculated with log rank test. RESULTS: A total of 21 male and 3 female with a mean age of 53.12 years were enrolled. Sixty-five percent of these received three cycles of chemotherapy. Five patients were positive for HPV 16 and none for HPV 18. Twenty-two of 24 patients showed GADD45 expression, 3 showed expression of Her-2-Neu while all 24 showed expression for EGFR1 protein. Two-year overall survival was 81%; GADD45 expressions were found to significantly affect the overall and disease-free survival, while any of the other protein expression studied and HPV status was not significant. CONCLUSION: The result of the present study shows significant downgrading of the oral cancers with neoadjuvant chemotherapy suggesting its utility in borderline operable cases. However, the response of chemotherapy does not appear to be related to the expression of EGFR, Her-2-Neu, and GADD45 protein or presence of HPV. Bone involvement, perineural invasion, and GADD45 expression significantly predict OS and DFS. All patients who did not express Gadd45 died before 2 years. Study with more subjects and longer follow-up should be carried out to elucidate this relation further.

Expression Analysis of Survivin and XIAP in Gallbladder Cancer: a Case-control Study in Indo-Gangetic Plain.

PURPOSE: Gallbladder cancer is a highly mortal disease with poor prognosis because of late presentation of disease. Survivin and X-linked inhibitor of apoptosis (XIAP) are one of the two important members of inhibitors of apoptosis. Thus, this study aimed to look at the expression of Survivin and XIAP in gallbladder cancer patients. METHODS: Survivin and XIAP expression were investigated in tissues of gallbladder cancer patients (40 cases) and compared with cholelithiasis as control (40 cases) by using immunohistochemistry. Their expression was correlated with clinicopathological parameters. RESULTS: Significantly higher (p < 0.05), Survivin protein was expressed in gallbladder cancer (n = 67.5%) than control (n = 35%). But it did not show any significant association with any of the clinicopathological parameter while XIAP was not expressed in the GBC patients (p > 0.05). CONCLUSION: Overexpression of Survivin in gallbladder cancer suggests its possible role and association with poor prognosis. But XIAP has not been found to be associated with gallbladder carcinogenesis.

Comparative Analysis of Mutational Profile of Sonic hedgehog Gene in Gallbladder Cancer.

BACKGROUND: Gallbladder cancer has high incidence in northeastern India; mortality too is high as the disease is often diagnosed late. Numerous studies have shown the role of sonic hedgehog (shh) in different cancers, an important ligand of the hedgehog signaling pathway. AIM: This study was carried out to evaluate the shh gene mutations in gallbladder cancer patients. METHODS: PCR-SSCP was performed for shh gene in 50 samples each of gallbladder cancer, cholelithiasis, and control. The samples showing aberration in banding pattern were sequenced. RESULTS: Variation in banding pattern was observed in 20% gallbladder cancer cases, 10% in cholelithiasis, and none of the control (χ 2 = 11.111; p < 0.05). Sequencing results revealed seven novel point mutations in GBC cases. These novel mutations were found to be associated with histopathology (p < 0.05) and stage (p < 0.05) of gallbladder cancer. CONCLUSION: This study reveals several novel individual and repetitive mutations of shh gene in GBC and cholelithiasis samples that may be used as diagnostic markers for gallbladder carcinogenesis.

Association of Methylenetetrahydrafolate Reductase Gene Polymorphism (MTHFR) in Patients with Gallbladder Cancer.

PURPOSE: 5,10-Methylenetetrahydrofolate reductase (MTHFR) is a key enzyme in folate metabolism and plays a major role in DNA methylation. There are two popular MTHFR polymorphisms known as C677T and A1298C which are found to be involved in folate metabolism and lowering the enzyme activity, thus may be linked with cancer development. This study aims to look at the association of these polymorphisms in gallbladder cancer. METHODS: Thirty patients each with gallbladder cancer, cholelithiasis, and normal gallbladder were genotyped for the above-given polymorphisms by PCR-restriction fragment length polymorphism (RFLP) method. RESULTS: C677T MTHFR polymorphism was not associated (χ(2) = 2.44, p = 0.85) with an increased likelihood of having gallbladder cancer. A1298C was significantly associated (χ(2) = 28.87, p < 0.001) with risk of developing gallbladder cancer. A1298C was significantly correlated with grade (r = 0.337, p < 0.001) and histopathology (r = 0.446, p < 0.001). CONCLUSION: This study proposed that MTHFR A1298C polymorphism may be associated with risk of developing gallbladder cancer, and there is no association between C677T polymorphism and gallbladder cancer.

Gall bladder carcinoma: Aggressive malignancy with protean loco-regional and distant spread.

The most common malignancy of biliary tract is gallbladder cancer (GBC) which is the third most common cancer in gastrointestinal tract. It is a lethal disease for most patients in spite of growing awareness and improved diagnostic techniques. GBC has a very poor prognosis and the 5 year survival rate is < 10%. Although etiology of the carcinoma of the gallbladder is still obscure, various factors have been implicated, cholelithiasis being the most frequent. The incidence of GBC worldwide is based on the gender, geography and ethnicity which suggest that both genetic and environmental factors can cause GBC. The major route of spread of gallbladder cancer (GC) is loco-regional rather than distant. It spreads by lymphatic, vascular, neural, intraperitoneal, and intraductal routes. Sonography is usually the most common imaging test to evaluate symptoms of biliary tract disease including suspected GC. With recent advances in imaging modalities like multi-detector computed tomography (CT) scanners, magnetic resonance imaging-positron emission tomography/CT diagnosis of gallbladder cancer has improved. Studies have also targeted molecular and genetic pathways. Treatment options have included extended and radical surgeries and adjuvant chemotherapy. This review article deals in detail with important aspects of carcinoma gallbladder and its manifestations and challenges. Role of various imaging modalities in characterization and accurate staging has been discussed. The loco-regional spread of this aggressive malignancy is dealt explicitly.

Association of mustard oil as cooking media with carcinoma of the gallbladder.

PURPOSE: Carcinoma of the gallbladder (CaGB) is a common health problem in Northern India. Exact causative factors are still obscure. Dietary habits are also known to be a major factor in the gallbladder carcinogenesis. Mustard oil is mostly used as cooking media, which is adulterated by sanguinarine, diethylnitrosamine and repeated frying. We tried to find out the association of mustard oil as cooking media with CaGB. METHODS: Twenty patients each of CaGB (group I) and cholelithiasis (group II) were included in the study. Sanguinarine and diethylnitrosamine (DEN) were extracted from the tissue and blood samples from both groups. Mean and standard error of mean of the concentration of the sanguinarine and DEN were calculated. Mann-Whitney U test was applied to test the level of significance between the two groups. RESULTS: The mean concentration of tissue sanguinarine in both groups (I and II) was 195.18 ng/mg and 24.05 ng/mg, respectively, and the difference was statistically highly significant (p < 0.001). The estimated concentration of blood sanguinarine was 230.96 ng/mL and 14.0 ng/mL in group I and II, respectively, and the difference was statistically highly significant (p < 0.001). The concentration of DEN in the tissue sample was 38.08 ng/mg in CaGB and 2.51 ng/mg in cholelithiasis patient, and these values were statistically highly significant (p < 0.001). Similarly, blood DEN concentration was 119.05 ng/mL and 4.22 ng/mL in group I and II, respectively, and the difference was statistically highly significant (p < 0.001). CONCLUSION: There is an increase in concentration of sanguinarine and diethylnitrosamine in CaGB blood and tissue in comparison to the cholelithiasis group suggesting an association with carcinoma of the gallbladder.

Chromosomal structural analysis in carcinoma of the gallbladder.

BACKGROUND: The etiopathogenesis of gallbladder cancer is still unknown. Both environmental and patient factors have been incriminated in its cause. That it is found in pockets of epidemiological distribution raises an issue of genetic changes associated with it. The aim of this study was to find out the chromosomal changes in gallbladder cancer. METHODS: Lymphocyte cell culture was carried out on blood of gallbladder cancer patients to determine chromosomal banding abnormalities. Native PAGE was also evaluated to analyze lactate dehydrogenase (LDH), superoxide dismutase (SOD) and catalase enzyme activity from the same blood of gallbladder cancer patients. RESULTS: Out of 30 gallbladder cancer patients, 4 male showed breakage on the long arm of chromosome 1 while only one male patient showed the translocation from the long arm of chromosome 4 to the long arm of chromosome 6 in a male patient. CONCLUSION: The aberrations found in our study may suggest underlying genetic predisposition for the development of gallbladder cancer. They can act as a marker for gallbladder cancer, which needs further study.

Helicobacter bilis in human gallbladder cancer: results of a case-control study and a meta-analysis.

INTRODUCTION: Gallbladder cancer is an uncommon neoplasm of uncertain etiology and poor survival. Recently, interest has been generated in bacterial infections and cancers. Helicobacter is one such bacterium found to be associated with gastric MALToma, gastric adenocarcinoma and hepatobiliary neoplasms. PATIENTS AND METHODS: Fifty four gallbladder cancer and 55 controls with cholelithiasis were studied. Helicobacter bilis was identified using 16S rRNA PCR. Relative risk and odds ratio with 95% CI were estimated. A detailed search of literature was carried out and selected relevant articles were extracted. A meta analysis was carried out using a random effect model. RESULTS: Helicobacter bilis was identified in 32/54 patients and 32/55 controls, The relative risk of gallbladder cancer in H. bilis positive cases was 1.05 (95% CI 0.49 to 2.24). Of the 10 identified case control studies on Helicobacter in the hepatobiliary tract 3 each were on gallbladder cancer and H. bilis. In meta analysis a pooled odds ratio of 4.13 (95% CI 2.68-6.36) favoring Helicobacter was observed. Pooled analysis of published studies on gallbladder cancer showed an odds ratio of 1.24 (95% CI 0.63-2.44). CONCLUSIONS: The present study failed to demonstrate any increase in risk of gallbladder cancer in presence of Helicobacter bilis. It may be hypothesized that increased risk observed in earlier studies may be indirectly due to increase in the risk of gallstones, although lack of any study specifically looking at this aspect and absence of normal controls in the present study makes this assumption superfluous.