Utility of guided FNAC and cell block preparation from liver and gall bladder masses: Learning experience from a tertiary care center.

INTRODUCTION: Ultrasound- and CT-guided fine needle aspiration cytology (FNAC) increases the accessibility of intra-abdominal masses to the liver and gall bladder with the advantages of low cost and high diagnostic yield. Cell block technique has been known for further increasing the diagnostic accuracy. AIMS AND OBJECTIVES: We aimed to study the effectiveness of FNAC and the cell block method in cytological diagnosis of liver and gall bladder masses. We also followed a step-wise approach to increase the success rate. MATERIALS AND METHODS: A 2-year observational study was done from July 2020 to June 2022. Total 80 guided (CT and ultrasound) aspirations were done from space occupying/mass lesions in the liver [74 (92.5%)] and gall bladder [6 (7.5%)], out of which cell blocks by the plasma thrombin method were prepared in 12 cases (15%). The on-site radiological details were noted, and rapid on-site evaluation was done in 65 cases (81.25%). The prepared cytology slides were stained with Papanicolaou, H and E and May-Grunwald Giemsa (MGG) stain. The cytological diagnosis was noted, and the uses and limitations (if any) were observed in each case. A step-wise structured questionnaire format was developed to assist the reporting pathologist so as not to miss out on important diagnostic observations, if present. RESULTS: FNAC in 71 cases (88.7%) gave a conclusive diagnosis. The maximum number of cases were of adenocarcinoma [38 (51.3%)] from the liver followed by hepatocellular carcinoma in 10 cases (13.5%). In gall bladder masses, all 6 cases (100%) were positive for malignancy, out of which 4 cases (66.7%) could be characterized as adenocarcinoma. The cell block preparation was helpful in reaching the diagnosis as well as typing the malignancy in 10 cases (83.3%). The chief limitation observed on conventional cytology smears was inadequate cellularity, which caused inconclusive diagnosis in 9 cases (11.25%). The reporting questionnaire was helpful chiefly in terms of time-efficient reporting in 34 cases (42.5%), increasing the ease and confidence in 69 cases (86.25%) and the advantage of reproducibility of data in all cases (100%) according to the case-by-case evaluation by the reporting pathologists. CONCLUSION: Guided FNAC in conjunction with the cell block technique is extremely helpful in the evaluation of mass lesions of the liver and gall bladder for cytological diagnosis. A proper step-wise approach may be useful to reach a quick and effective diagnosis.

PE_PGRS antigens of Mycobacterium tuberculosis induce maturation and activation of human dendritic cells.

Mycobacterium tuberculosis, the causative agent of pulmonary tuberculosis, infects one-third of the world's population. Activation of host immune responses for containment of mycobacterial infections involves participation of innate immune cells, such as dendritic cells (DCs). DCs are sentinels of the immune system and are important for eliciting both primary and secondary immune responses to pathogens. In this context, to understand the molecular pathogenesis of tuberculosis and host response to mycobacteria and to conceive prospective vaccine candidates, it is important to understand how cell wall Ags of M. tuberculosis and, in particular, the proline-glutamic acid_polymorphic guanine-cytosine-rich sequence (PE_PGRS) family of proteins modulate DC maturation and function. In this study, we demonstrate that two cell wall-associated/secretory PE_PGRS proteins, PE_PGRS 17 (Rv0978c) and PE_PGRS 11 (Rv0754), recognize TLR2, induce maturation and activation of human DCs, and enhance the ability of DCs to stimulate CD4(+) T cells. We further found that PE_PGRS protein-mediated activation of DCs involves participation of ERK1/2, p38 MAPK, and NF-kappaB signaling pathways. Priming of human DCs with IFN-gamma further augmented PE_PGRS 17 or PE_PGRS 11 Ag-induced DC maturation and secretion of key proinflammatory cytokines. Our results suggest that by activating DCs, PE_PGRS proteins, important mycobacterial cell wall Ags, could potentially contribute in the initiation of innate immune responses during tuberculosis infection and hence regulate the clinical course of tuberculosis.

The multifunctional PE_PGRS11 protein from Mycobacterium tuberculosis plays a role in regulating resistance to oxidative stress.

Mycobacterium tuberculosis utilizes unique strategies to survive amid the hostile environment of infected host cells. Infection-specific expression of a unique mycobacterial cell surface antigen that could modulate key signaling cascades can act as a key survival strategy in curtailing host effector responses like oxidative stress. We demonstrate here that hypothetical PE_PGRS11 ORF encodes a functional phosphoglycerate mutase. The transcriptional analysis revealed that PE_PGRS11 is a hypoxia-responsive gene, and enforced expression of PE_PGRS11 by recombinant adenovirus or Mycobacterium smegmatis imparted resistance to alveolar epithelial cells against oxidative stress. PE_PGRS11-induced resistance to oxidative stress necessitated the modulation of genetic signatures like induced expression of Bcl2 or COX-2. This modulation of specific antiapoptotic molecular signatures involved recognition of PE_PGRS11 by TLR2 and subsequent activation of the PI3K-ERK1/2-NF-κB signaling axis. Furthermore, PE_PGRS11 markedly diminished H(2)O(2)-induced p38 MAPK activation. Interestingly, PE_PGRS11 protein was exposed at the mycobacterial cell surface and was involved in survival of mycobacteria under oxidative stress. Furthermore, PE_PGRS11 displayed differential B cell responses during tuberculosis infection. Taken together, our investigation identified PE_PGRS11 as an in vivo expressed immunodominant antigen that plays a crucial role in modulating cellular life span restrictions imposed during oxidative stress by triggering TLR2-dependent expression of COX-2 and Bcl2. These observations clearly provide a mechanistic basis for the rescue of pathogenic Mycobacterium-infected lung epithelial cells from oxidative stress.

Resveratrol as an adjunct therapy in cyclophosphamide-treated MCF-7 cells and breast tumor explants.

Cyclophosphamide (CPA) has efficacy as a breast cancer therapy. However, toxicity to CPA limits its clinical applications. Hence there is a need to develop compounds that may be combined with it to improve the efficacy and overcome toxicity. We showed previously that Resveratrol (RES), a chemopreventive agent, increased the growth inhibitory effect of CPA-treated MCF-7 cells. Here we have explored the molecular basis of 5 mM CPA and 50 µM RES as a combination on cell-cycle progression, apoptosis and oxidative stress in MCF-7 breast cancer cells. Efficacy of the combination was also evaluated in a serum-free tumor explant culture model. The combination elicited enhanced anti-proliferative action coupled with differential expression of cell-cycle, apoptosis and stress factors. Furthermore, co-treatment superiority in histologically validated ER positive breast cancer explants suggests that this combination may be a worthy future clinical anti-neoplastic regimen.

Protective association exhibited by the single nucleotide polymorphism (SNP) rs1052133 in the gene human 8-oxoguanine DNA glycosylase (hOGG1) with the risk of squamous cell carcinomas of the head & neck (SCCHN) among north Indians.

BACKGROUND & OBJECTIVES: Imbalances in compactly regulated DNA repair pathways in the form of single nucleotide polymorphisms (SNPs) within vital DNA repair genes may result in insufficient DNA repair and increase in DNA breaks thus rendering the human system vulnerable to the debilitatory effects of grave diseases like cancers. The present study involves investigation of association of the non-synonymous SNP rs1052133 (C8069G/Ser326Cys) located in the exonic region of the gene human 8-oxoguanine DNA glycosylase (hOGG1) with the risk of squamous cell carcinomas of the head and neck (SCCHN). METHODS: Case-control based genetic association study was performed among 575 (250 SCCHN cases and 325 normal healthy controls) sub-population cluster-matched (Indo-Europeans linguistic subgroup + Caucasoid morphological subtype) samples from the north Indian States of Uttar Pradesh and Uttarakhand using polymerase chain reaction followed by restriction fragment length polymorphism (PCR-RFLP) and DNA sequencing analysis. RESULTS: Our results demonstrated statistically significant protective association for the heterozygous CG [Odds Ratio (OR) 0.6587, 95% Confidence Interval (CI) 0.4615 to 0.9402, P=0.0238], homozygous mutant GG (OR 0.2570, 95% CI 0.1070 to 0.6175, P=0.0013) and combined mutant CG + GG (OR 0.6057, 95% CI 0.4272 to 0.8586, P=0.0059) genotypes. INTERPRETATION & CONCLUSIONS: The results indicate that the polymorphism rs1052133 is strongly associated with SCCHN susceptibility and the mutant (G) allele might be a protective factor for SCCHN among north Indian subpopulations.

A quantitative and qualitative comparative analysis of collagen fibers to determine the role of connective tissue stroma in oral squamous cell carcinoma using special stains and polarized microscopy.

BACKGROUND: Solid tumors such as oral squamous cell carcinoma (OSCC) are composed of malignant epithelial cells and the stroma in which these cells are dispersed. As the tumor progresses, the extracellular matrix undergoes dramatic morphological and architectural changes. Special stains make analysis easy and less erroneous by highlighting the area of interest and can be used to study these changes. AIM: The aim of the study was to analyze morphological changes in collagen fibers in various histological grades of OSCC using Masson's trichrome (MT) and Picrosirius red (PSR). STUDY DESIGN: The study comprised 74 tissue samples, divided into two groups: Group I consisted of 63 cases of histologically proven OSCC (39 cases of well-differentiated squamous cell carcinoma [WDSCC], 17 moderately differentiated squamous cell carcinoma [MDSCC] and 7 poorly differentiated squamous cell carcinoma [PDSCC]) and Group II consisted of 11 cases of normal mucosa as controls. MATERIALS AND METHODS: Sections were stained with hematoxylin and eosin, MT and PSR and observed under light and polarizing microscope, respectively. STATISTICAL ANALYSIS: ANOVA, Tukey's honestly significant difference post hoc multiple comparison test, Chi-square test and paired t-test were used for the statistical analysis. RESULTS: As the grade of OSCC progressed, collagen fibers became thin, loosely packed and haphazard. The mean area fraction also decreased. They exhibited orange-red hue and strong birefringence in WDSCC, yellowish-orange hue and strong birefringence in MDSCC and greenish-yellow hue and weak birefringence in PDSCC. CONCLUSION: Initially, there is a reorganization of the collagen fibers in an attempt to prevent the invasion of tumor cells, but as cancer progresses, the stromal change enhances movement of the tumor cells within it, leading to metastasis.

Association of polymorphisms in base excision repair genes with the risk of breast cancer: a case-control study in North Indian women.

Inheritance of common genetic variants at one or more base excision repair (BER) genes may result in a reduced DNA repair capacity and in an increased risk of cancers like breast cancer. The present case-control study with 390 north Indian women (155 breast cancer cases and 235 controls) was aimed to investigate the association of seven nonsynonymous BER gene polymorphisms viz. rs1130409/T1865G (APEX1), rs1799782/T22142C (XRCC1), rs25487/G23990A (XRCC1), rs4989588/T3337A (FEN1), rs4989586/ G3259A (FEN1), rs4989587/C3315T (FEN1), and rs1050525/G6941T (PCNA) with breast cancer susceptibility. Statistically significant association with breast cancer risk was observed for rs1130409 homozygous mutant GG [odds ratio (OR) 3.35, 95% confidence interval (CI) 1.36-8.26), heterozygous GT (OR 2.42, 95% CI 1.56-3.76), and combined mutant (GT + GG) (OR 2.52, 95% CI 1.65-3.86] genotypes and rs25487 homozygous mutant AA (OR 2.91, 95% CI 1.66-5.10) and combined mutant (AA + AG) (OR 1.41, 95% CI 0.903-2.19) genotypes, whereas protective association was exhibited by rs1799782 homozygous mutant CC (OR 0.413, 95% CI 0.082-2.08), heterozygous TC (OR 0.351, 95% CI 0.189-0.650), and combined mutant (TC + CC) (OR 0.357, 95% CI 0.199-0.641) genotypes. Association study using reconstructed haplotypes of XRCC1 gene showed positive association for the TA haplotype (OR 2.014, 95% CI 1.462-2.775) and a protective association for the CG haplotype (OR 0.173, 95% CI 0.052-0.576) pertaining to breast cancer risk. The results indicate that the polymorphisms rs1130409 (APEX1) and rs25487 (XRCC1) might be involved in contributing towards breast cancer susceptibility, while rs1799782 (XRCC1) might have protective influence.

Apoptosis triggered by Rv1818c, a PE family gene from Mycobacterium tuberculosis is regulated by mitochondrial intermediates in T cells.

Ectopic expression of the Mycobacterium tuberculosis PE-family gene Rv1818c, triggers apoptosis in the mammalian Jurkat T cells, which is blocked by anti-apoptotic protein Bcl-2. Although complete overlap is not observed, a considerable proportion of cellular pools of ectopically expressed Rv1818c localizes to mitochondria. However, recombinant Rv1818c does not trigger release of cytochrome c from isolated mitochondria even though Rv1818c protein induced apoptosis of Jurkat T cells. Apoptosis induced by Rv1818c is blocked by the broad-spectrum caspase inhibitory peptide zVAD-FMK. Unexpectedly, Rv1818c-induced apoptosis is not blocked in a Jurkat sub-clone deficient for caspase-8 (JI 9.2) or in cells where caspase-9 function is inhibited or expression of caspase-9 reduced by siRNA, arguing against a central role for these caspases in Rv1818c-induced apoptotic signaling. Depleting cellular pools of the mitochondrial protein Smac/DIABLO substantially reduces apoptosis consistent with mitochondrial involvement in this death pathway. We present evidence that Rv1818c-induced apoptosis is blocked by the co-transfection of an endogenous inhibitor of caspase activation, XIAP in T cells. Additionally, Rv1818c is released into extracellular environment via exosomes secreted by M. tuberculosis infected BM-DC's and macrophages. Furthermore, the extracellular Rv1818c protein can be detected in T cells co-cultured with infected BM-DC's. Taken together, these data suggest that Rv1818c-induced apoptotic signaling is likely regulated in part by the Smac-dependent activation of caspases in T cells.

Association of CYP1A1 polymorphisms with breast cancer in North Indian women.

Cytochrome P-450 (CYP) 1A1 is a candidate gene for low penetrance breast cancer (BC) susceptibility. Evidences demonstrate that ethnic differences in BC incidence may be partly due to genetic factors, including polymorphisms in the genes. In the present case control study four CYP1A1 gene polymorphisms, m1 (T6235C), m2 (A4889G), m3 (T5639C), and m4 (C4887A) were studied for their association with BC conjointly with the known risk factors such as age, menopausal status, diet, and life style. Polymorphisms of CYP1A1 gene were detected by PCR-RFLP method. The homozygous mutant (G/G) of m2 polymorphism was significantly associated with BC. Consequently, association of both m2 heterozygous mutant genotype (A/G) and combined group [homozygous (G/G) plus heterozygous (A/G) mutant genotype] showed association with postmenopausal women. Incidences of BC were also found to be independent of clinicopathological factors except heterozygous mutant genotype (A/G) m2 showed association with dietary factors and high grade tumors while homozygous mutant (G/G) m2 showed association with ER/PR-positive BC cases. Wild-type m3 was observed in all the subjects in cases as well as in controls. No significant association was observed between m1 and m3 polymorphisms and BC risk in all the subjects as well as when stratified into pre- and postmenopausal subjects. This indicates that out of ml and m2 polymorphisms that have been reported in Asians, only m2 is associated with North Indians.

Massive ovarian edema--a diagnostic dilemma: a case report.

A case of massive ovarian edema and polycystic ovaries in a 15 years old girl is being reported for its rarity. Definitive preoperative diagnosis is often not achieved and there by the patient is at a risk of losing the ovary(ies). Thus a definitive preoperative diagnosis should be ascertained to save the organ. A review on the previously reported cases with probable etiopathogenesis and emphasis on the radiological and morphological diagnosis for appropriate management has been discussed.

Securing the food supply chain: understanding complex interdependence through agent-based simulation.

The food industry has many points of vulnerability in its supply chain. It currently lacks integrated crisis management and response programs to understand the importance of decision-making during and in the aftermath of a bioterrorist attack on the food supply. Computer simulations have been used successfully in other industries as training and analysis tools. This paper describes an agent-based simulation for food defense training and analysis. Production information, consumption patterns, morbidity/mortality rates, recall costs and additional information were collected and provided to a data-driven simulation to anticipate the impact of decision-making on economic and public health during a terrorist attack. A case study is given with a representative exercise involving forty industry representatives who participated in a food defense simulation. Their decisions (recall and microbiological and toxicological testing) were derived from testing results, press releases, epidemiological data, and discussions with other industry and regulatory teams. Decisions made during the simulation resulted in over 76,000 illnesses, 45 deaths, and $132 million in recall costs. The no intervention, baseline scenario estimated to result in 91,000 illnesses and 54 deaths, indicating the improved public health outcomes resulting from players' decisions. Participants identified three key learning points: 1) communication between all groups is pertinent and challenging, 2) approaches to solve inherent food safety problems cannot be used to address food defense situations, and 3) human resource procedures regarding new hires and disgruntled employees should involve additional security measures. This computer simulation could be a valuable resource in food defense awareness and help educate companies and regulators about food defense risks and decision-making consequences.

Extra osseous osteosarcoma of the retroperitoneum: an unusual entity.

Extra-osseous osteosarcomas constitute about 1-1.2% of all osteosarcomas. The most common sites are the extremities, thorax, and the abdomen. Retroperitoneal osteosarcomas are rare and very few cases have been reported. They are similar in their biology to high grade soft tissue sarcomas. R0 resection appears to be the best possible treatment for these tumors. All three variants of conventional osteosarcoma--osteoblastic, chondroblastic, and fibroblastic have been described in these tumors. Chemotherapy has been attempted with adriamycin-based regimens with poor results. Unlike extremity osteosarcomas, these tumors have been found to be chemoresistant. The 5 year survival has ranged from a dismal 12% to about 25%. We report a 46-year-old male who presented with a kidney tumor infiltrating the descending colon, but turned out to be an extra osseous osteosarcoma. An R0 resection was done and adjuvant chemotherapy given.

Statistically significant association of the single nucleotide polymorphism (SNP) rs13181 (ERCC2) with predisposition to Squamous Cell Carcinomas of the Head and Neck (SCCHN) and Breast cancer in the north Indian population.

BACKGROUND: Non-synonymous single nucleotide polymorphisms (SNPs) within vital DNA repair genes may cause reduction of activity leaving the genome unrepaired resulting in genomic instability and cancer. MATERIALS AND METHODS: The present endeavour involved study on the association of the SNP rs13181 (Lys751Gln/A18911C) in the Nucleotide Excision Repair (NER) pathway gene ERCC2 (excision repair cross-complementing rodent repair deficiency, complementation group 2) with the risks of Squamous Cell Carcinomas of the Head and Neck (SCCHN) and Breast cancer using a case-control based association study among 685 (400 controls and 285 SCCHN-affected cases) and 395 (227 normal healthy female controls and 168 breast cancer cases) ethnically-matched samples, respectively from north India using Polymerase Chain Reaction followed by Restriction Fragment Length Polymorphism (PCR-RFLP) analysis. RESULTS: Results showed significant association of rs13181 homozygous mutant (CC) [Odds Ratio (OR) 4.412, 95% Confidence Interval (CI) 2.413 to 8.068], heterozygous (AC) (OR 2.086, 95% CI 1.246 to 3.492) and combined mutant (AC + CC) (OR 2.672, 95% CI 1.647 to 4.334) genotypes with predisposition to Breast cancer. Statistically significant increase in SCCHN risk was also associated with the mutant genotypes of rs13181 (ERCC2), viz. homozygous mutant (CC) (OR 1.680, 95% CI 1.014 to 2.784), heterozygous (AC) (OR 1.531, 95% CI 1.092 to 2.149) and combined mutant (AC + CC) (OR 1.560, 95% CI 1.128 to 2.158) genotypes. CONCLUSION: The results of this case-control study indicate that the polymorphism rs13181 might be a risk factor for predisposition towards SCCHN and breast cancer among north Indian subpopulations.