Clinical Significance of Overexpression of Oct4 in Advanced Stage Gallbladder Carcinoma.

BACKGROUND: Oct4 has critical role in maintaining pluripotency, proliferative potential, and self-renewal capacity in embryonic stem and germ cells. Although Oct4 has been shown to be upregulated in many cancers, its clinical significance in gallbladder carcinoma is poorly understood. METHODS: We studied the expression profile of Oct4 in 61 GBC and 30 chronic cholecystitis (as control) using real time RT-PCR, western blotting, and immunohistochemistry. The expression data was correlated with clinico-pathological parameters. The diagnostic utility was assessed through ROC curve, and prognostic value was analyzed by Kaplan-Meier method. RESULTS: Oct4 was significantly upregulated at mRNA as well as protein levels. The higher mRNA expression shows significant association with late stage, late T stage, and higher grade of tumor. A significant positive correlation was also observed with stage, T stage, and tumor grade. Sum score analysis of protein expression shows positive correlation with stage and the presence or absence of gallstone in tumor samples. The ROC curve analysis revealed the moderate diagnostic potential of Oct4. Kaplan-Meier analysis showed that patients having higher expression of Oct4 were having low mean survival compared with the patients with lower Oct4 expression. CONCLUSION: In conclusion, our data suggests that higher expression of Oct4 may serve as potential biological indicator for tumor aggressiveness and poor prognosis of GBC.

Association of haplotype and linkage disequilibrium of PARP1 polymorphisms rs1136410, rs1805405 and rs3219088 with gallbladder cancer.

BACKGROUND: Previously, we have reported that PARP1 rs1136410 is significantly associated with increased the risk of gallbladder cancer. AIM: We aimed to investigate the association of PARP1 rs1805405 and rs3219088 polymorphisms with risk of GBC and also association of the haplotype and combined effect of PARP1 SNPs (rs1805405 G/A, rs3219088 G/T and rs1136410 A/G). We have also investigated the expression profile of PARP1 and its correlation with polymorphisms, clinical parameters and overall survival. METHODS: PARP1 polymorphisms were genotyped by PCR-RFLP and the expression profile of PARP1 at mRNA level was analyzed by semi-quantitative PCR. Overall survival was analyzed using Kaplan-Meier plot and Cox-regression analysis. RESULTS: Haplotype analysis of the PARP1 polymorphisms revealed that AGG, AAG and GGT haplotypes are significantly associated with decreased risk of GBC, while AAT, AGT, GGG and GAG haplotypes are significantly associated with increased risk of GBC. Patients with T1+T2 and treated with chemotherapy having risk genotypes of rs1805405 have decreased overall survival. Upregulation of PARP1 is significantly associated with longer overall survival in patients with GBC with different clinical parameters. SNPs rs1136410 and rs1805405 genotypes are significantly associated with PARP1 expression. CONCLUSION: Haplotype analysis suggests that PARP1 may have a potential role in gallbladder carcinogenesis.

Frequent promoter hypermethylation and down regulation of BNIP3: An early event during gallbladder cancer progression.

BACKGROUND: Epigenetic alterations have been reported as one of the risk factors of gallbladder cancer. Promoter hypermethylation is associated with high incidence and poor prognosis of GBC. Bcl-2/adenovirus E1B 19 kDa interacting protein 3 is a pro-apoptotic protein member of Bcl-2 family. AIMS: Present study was aimed to investigate expression profile and promoter methylation status of BNIP3 in GBC and its correlation with clinico-pathological parameters. METHODS: The expression analysis and methylation status of BNIP3 was performed by semi-quantitative reverse transcription polymerase chain reaction and Methylation-specific polymerase chain reaction respectively in 84 GBC patients and 29 gallstone tissues (used as normal controls). RESULTS: We demonstrate down regulation of BNIP3 in 56% of the GBC samples. BNIP3 promoter is also frequently hypermethylated (69%) in GBC samples. Interestingly, we found that 69% (40/58) of the BNIP3 promoter hypermethylated samples had also reduced expression of BNIP3. Our data demonstrate significant correlation of the mRNA expression and promoter hypermethylation with late stage and nodal metastasis. Hypermethylation of BNIP3 promoter is associated with low overall survival period. CONCLUSION: Our results suggest that promoter hypermethylation is an early event and can be a frequent mechanism for downregulation of BNIP3 in GBC.

Expression of Poly(Adenosine Diphosphate-Ribose) Polymerase Protein in Breast Cancer.

Background: The use of poly(adenosine diphosphate-ribose) polymerase (PARP) inhibitors for breast cancer (BC) therapy is the subject of debate, and there is an urgent need to understand much the expression and prognostic role of the PARP1 protein. In this study, we have compared the expression of PARP between BC and benign breast disease (BBD) patients and also analyzed the association of PARP expression with clinicopathological parameters in BC. Methods: The study consists of 30 patients with newly diagnosed operable BC who were planned for surgery without neoadjuvant chemotherapy and 15 patients of BBD as a control between 2019 and 2021. Immunohistochemical analyses were performed prospectively on tissue samples. Anti-human PARP1 rabbit polyclonal antibody gives strong nuclear positivity. Internal control was the adipose tissue and the BBD acted as the external control. PARP1 expression was evaluated using the multiplicative quickscore method. Results: The mean age for BC patients was 51.30 ± 10.694 years (range: 25-75 years) while BBD was below 30 years. Overexpression of PARP was present in 25 (83.3%) and weak expression in 5 (16.7%) of BC patients compared to BBD, only 2 (13.3%) patients demonstrated an overexpression of PARP, and 13 (86.6%) patients showed weak expression which showed significant association (P < 0.001). In BC, nuclear PARP (nPARP) overexpression was seen in 22 (73.3%) patients and weak expression of nPARP in 8 (26.7%), whereas 5 (16.7%) patients showed cytoplasmic overexpression. On comparing expression of PARP with clinicopathological parameters, PARP overexpression was significantly associated with older population (age >50 years) (P = 0.002), postmenopausal women (P = 0.029), higher TNM stage (Stage II and III) (P = 0.014), higher histological grade (grade 2) (P = 0.043), and presence of lymphovascular invasion (P = 0.015). Enhanced PARP1 expression is closely correlated with positive estrogen receptor status (P = 0.001) and PR status (P = 0.001). Overall PARP and nPARP overexpression was significantly associated with ER- (P = 0.006 and P = 0.008) and PR-positive (P = 0.006 and P = 0.008) patients. The PARP and nPARP overexpression was significantly associated with nontriple-negative BC patients (P = 0.001 and P = 0.001). Conclusion: We have not come across any study in the literature to compare PARP expression in BC and BBD patients. On the basis of our observations, we concluded that PARP overexpression is a poor prognostic marker in BC.

Frequent Downregulation and Promoter Hypermethylation of DLC1: Relationship with Clinical Outcome in Gallbladder Cancer.

INTRODUCTION: Down regulation of DLC1 is associated with poor prognosis in many cancers, however, its role in gallbladder cancer (GBC) is still unclear. In present study, we investigated the expression profile and promoter methylation status of DLC1. METHODS: Expression profiles of DLC1 in 55 GBC and their paired adjacent control samples were analyzed through real time RT-PCR and immunohistochemistry. The mRNA data was correlated with clinico-pathological parameters. Promoter hypermethylation was analyzed through MSP. RESULTS: DLC1 shows downregulation in 76.4%, upregulation in 10.9% whereas no change in 12.7% of GBC samples. Its underexpression shows significant correlation with tumor grade and nodal spread. IHC shows cytoplasmic expression of DLC1 in normal as well as tumor samples. IHC result was concordant to mRNA result. Samples having downregulated DLC1 expression show heterozygous methylation in 83.3% of samples and homozygous methylation in 9.5% of samples whereas 7% of samples have no methylation. Kaplan-Meier analysis shows patient with decreased mRNA of DLC1 have significant low mean survival compared to patients with higher mRNA expression of DLC1. CONCLUSION: Our findings suggested that dysregulated expression of DLC1 and its hypermethylation may be one of the events playing roles in tumorigenesis of GBC and may serve as a potential target for development of future GBC gene therapy.

A Clinicopathological Study to Assess the Role of Intralesional Sclerotherapy Following Propranolol Treatment in Infantile Hemangioma.

Context: As propranolol has emerged as first-line therapy for problematic infantile hemangioma, the number of non-responders and partial responders to propranolol therapy is also increasing. Aims: The study was conducted to evaluate the response of intralesional bleomycin, triamcinolone, and a combination of both as second line of treatment for the residual hemangioma following propranolol therapy. Settings and Design: A prospective comparative study was conducted in patients who were either non-responders or partial responders to previous propranolol treatment. Materials and Methods: The patients randomly received injection bleomycin, injection triamcinolone, and combination of both bleomycin and triamcinolone. The response to treatment was recorded clinically by using photographs. The pathological response was assessed by calculating pre-treatment and post-treatment microvessel density in biopsy of lesion from the non-cosmetic sites using immunohistochemistry. Statistical Analysis Used: χ2 test was used to test the association between the variables. The utility of microvessel diameter (MVD) in terms of clinical response to the therapy was predicted by using receiver operating characteristic (ROC) curve. Results: Out of the 134 patients, 42 received bleomycin and 44 received triamcinolone and were treated with a combination of both. The overall clinical response was better in the combination group compared with the bleomycin group (P = 0.018) and triamcinolone group (P = 0.0005), respectively, after 6 months of follow-up. There was no difference in clinical response between the triamcinolone and bleomycin groups. Change in MVD correlated with the clinical response. Conclusion: The combination of bleomycin and triamcinolone is effective and safe for the treatment of residual hemangioma.

TRAIL receptors are differentially regulated and clinically significant in gallbladder cancer.

Deregulation of the receptors of TNF-related apoptosis inducing ligand (TRAIL) has been reported in various cancers. In an effort to define the role of these receptors we profiled their expression in gallbladder cancer (GBC) and explored their clinical significance. Expression of TRAIL receptors' mRNA in GBC was analysed through reverse transcriptase polymerase chain reaction (RT-PCR), and protein through western blotting, immunohistochemistry and enzyme-linked immunosorbent assay (ELISA). mRNA data show frequent higher expression of TRAIL receptors in GBC samples. Death receptors DR4 and DR5 showed significant negative correlation with tumour stage, T stage and tumour grade; DcR1 transcript showed positive correlation with tumour stage, N stage, M stage and tumour grade. Similarly, IHC showed frequent positive staining for DR4, DR5 and DcR1in GBC samples. Cytoplasmic and nuclear DR4 protein showed negative correlation with T stage and tumour grade, whereas cytoplasmic DcR1 protein showed positive correlation with tumour stage and N stage. Nuclear DcR1 showed positive correlation with N stage. ELISA results showed significantly higher expression of secretory DcR1 in GBC patients. Kaplan-Meier analysis demonstrated significantly decreased mean survival of patients with positive staining of cytoplasmic DcR1. High level of death receptors identified the patients with early gallbladder cancer, whereas high DcR1 expression served as a prognostic factor for poor outcome.

MoS2-Modified Curcumin Nanostructures: The Novel Theranostic Hybrid Having Potent Antibacterial and Antibiofilm Activities against Multidrug-Resistant Hypervirulent Klebsiella pneumoniae.

The recent emergence of hypervirulent clinical variants of Klebsiella pneumoniae (hvKP) causing community-acquired, invasive, metastatic, life-threatening infections of lungs, pleura, prostate, bones, joints, kidneys, spleen, muscles, soft-tissues, skin, eyes, central nervous system (CNS) including extrahepatic abscesses, and primary bacteremia even in healthy individuals has posed stern challenges before the existing treatment modalities. There is therefore an urgent need to look for specific and effective therapeutic alternatives against the said bacterial infection or recurrence. A new type of MoS2-modified curcumin nanostructure has been developed and evaluated as a potential alternative for the treatment of multidrug-resistant isolates. The curcumin quantum particles have been fabricated with MoS2 via a seed-mediated hydrothermal method, and the resulting MoS2-modified curcumin nanostructures (MQCs) have been subsequently tested for their antibacterial and antibiofilm properties against hypervirulent multidrug-resistant Klebsiella pneumoniae isolates. In the present study, we found MQCs inhibiting the bacterial growth at a minimal concentration of 0.0156 µg/mL, while complete inhibition of bacterial growth was evinced at concentration 0.125 µg/mL. Besides, we also investigated their biocompatibility both in vitro and in vivo. MQCs were found to be nontoxic to the SiHa cells at a dose as high as 1024 µg/mL on the basis of the tested adhesion, spreading of the cells, and also on the various serological, biochemical, and histological investigations of the vital organs and blood of the Charles Foster Rat. These results suggest that MQCs have potent antimicrobial activities against hvKP and other drug resistant isolates and therefore may be used as broad spectrum antibacterial and antibiofilm agents.

Effect of Agonist and Antagonist on the In Vitro Contractility of Inflamed Vermiform Appendix.

INTRODUCTION: Appendicitis poses a great health problem worldwide. Previous studies demonstrated structural damage to neuronal network and interstitial cell of Cajal in appendicitis. Above observations suggest for the alterations in appendicular motility/contractility in appendicitis. But the mechanisms involved in mediating the contractility in inflamed vermiform appendix is not known till date. AIM: The present in vitro study was performed to find out the mechanisms responsible for contractility in the inflamed human vermiform appendix. MATERIALS AND METHODS: Contractions of the longitudinal muscle strips of inflamed appendix were recorded in vitro at 37±0.5°C. Control contractions were recorded for 30 min after an initial tension of 0.5 gram. Initially dose-response experiments of agonists (acetylcholine, serotonin and histamine) were performed separately and the dose that produced maximum contraction was determined with each agonist. This maximal dose of agonist was used to elicit contractions in next series of experiments before and after pre-treatment with appropriate antagonists like atropine, ondansetron (5-HT3 antagonist) and chlorpheniramine maleate respectively. RESULTS: Acetylcholine (ACh) and serotonin (5-HT) elicited maximum amplitude of contraction at 10 µM and 1 µM concentration respectively. These contractions were significantly blocked by prior exposure of muscle strips with atropine (100 µM) and ondansetron (10 µM). Histamine produced very low amplitude of contractions in comparison to ACh or 5-HT and did not exhibit dose-response relations. The histamine induced contractions were blocked by H1 antagonist chlorpheniramine maleate (100 µM). CONCLUSION: The observations suggested that the contractility of longitudinal muscle strips of inflamed vermiform appendix in human beings was predominantly mediated by muscarinic and serotonergic (5-HT3) mechanisms, whereas, histaminergic mechanisms played a minor role in mediating the contractility.

Epidemiological Study of Triple-Negative Breast Cancer Patients in North Indian Population: a Hospital-Based Study.

Triple-negative breast cancer (TNBC) accounts for 10-25% of all breast tumors. This makes it more difficult to treat, so triple-negative cancers often require targeted therapies. We studied the prevalence of TNBC in a hospital-based study and compared the clinicopathological characteristics of triple-negative and non-triple-negative breast tumors. One hundred three patients were included in the study that underwent modified radical mastectomy. The procedure of immunostaining was performed using formalin-fixed, paraffin-embedded tissue sections. These sections were stained immunohistochemically for ER, PR, and HER2 neu by using ready-to-use monoclonal antibody detection system with 3'-3' diaminobenzidine hydrochloride (DAB) as chromogen. Of all 103 patients, 35 (34%) were triple negative. The average age of patients of TNBC and non-TNBC group was found as 44.16 and 40.73 years, respectively. Patients of post-menopausal state were higher than premenopausal in TNBC (22/35; 62%) and non-TNBC groups (45/68; 66%). Further, TNBC patients reported at clinically early stages I and II (18/35; 51.4%) while non-TNBC patients predominantly reported at later stages III and IV (44/68; 64.7%). It was also observed that breast tumor size in majority of the patients in both groups lies between 2 to 5 cm (TNBC = 23/35; 65.7% and non-TNBC = 35/68; 51.5%). Lymph node metastases were present in 51.5% (18/35) cases in TNBC patients and 64.7% (44/68) cases in non-TNBC group. Despite the limitation of less number of breast cancer cases, we analyzed that TNBC tumors have aggressive clinical values than non-TNBC, though having no statistically significant difference between the prognostic clinical parameters of two groups.

Nested Multiplex (NMPCR) Detection of Human Papillomavirus (HPV) 16 and 18 in Pre-invasive Lesions and its Implication in Screening of Carcinoma Cervix (CaCx).

INTRODUCTION: Carcinoma cervix (CaCx) is a preventable disease and is caused by certain high risk Papillomaviruses. In the present study, our aim was to investigate the utility of Nested Multiplex Polymerase Chain Reaction (NMPCR) in detecting Human Papillomavirus (HPV) 16 and 18 in cervical scrapes/biopsy samples and to correlate with cervical cytology/ histopathology findings. METHODS: A total of 119 females were subjected for Papanicolaou smear examination of cervical scrapes and/or histopathological examination of cervical tissues. These samples were also subjected to nested multiplex PCR targeting HPV 16/ 18 specific E6/7 gene sequences. RESULTS: HPV 16/18 were detected in 33.6% (40/119) cases included in the study. The overall HPV 16/ 18 positivity among cases with Negative for Intraepithelial Lesion or Malignancy, Low grade Squamous Intraepithelial Lesion, and High grade Squamous Intraepithelial Lesion was observed to be 20.8%, 44%, and 66.7% respectively. Positivity for HPV 16 in cases with Squamous Cell Carcinoma (SCC) was found to be 80%. HPV positivity among subjects reported with reactive cellular changes, a sub category of Negative for Intraepithelial Lesion or Malignancy, was observed to be 26.6%. CONCLUSION: HPV 16 and 18 positivity in cases reported with different stages of pre invasive lesions of CaCx, particularly in the subcategory reactive cellular changes of Negative for Intraepithelial Lesion or Malignancy, indicates that NMPCR detection of HPV 16/ 18 may be used as a screening tool for CaCx in conjunction with Papanicolaou smear examination.

Isolated tuberculosis of the ampulla of vater masquerading as periampullary carcinoma: a case report.

CONTEXT: Isolated tuberculosis of the ampulla of Vater has not yet been reported. The clinical features of isolated periampullary tuberculosis are at times similar to those seen in patients with periampullary carcinoma. Diagnosis is difficult, and biopsy and culture of the suspected lesion are often negative for Mycobacterium tuberculosis. CASE REPORT: We herein describe one such case masquerading as periampullary carcinoma in a 70-year-old woman. Due to comorbid conditions only a local excision of the ampulla was carried out. Histopathology revealed giant cells in the absence of caseation necrosis and the presence of Mycobacterium tuberculosis was proven using the polymerase chain reaction. CONCLUSION: Isolated tuberculosis of ampulla of Vater is extremely rare but must be kept in mind when making the differential diagnosis of isolated ampullary lesion.