To document the expression of activated forms of caspase-3 in human granulosa cells.
Laboratory study.
In vitro fertilization (IVF) laboratory of the Split University Hospital and laboratory of the ...Department of Anatomy, Histology, and Embryology.
Ovarian tissues were obtained from women undergoing hysterectomy/ovariectomy for benign conditions and human granulosa cells were obtained from women undergoing oocyte retrieval for IVF.
Immunostaining of tissue sections and cell smears using antibody to active caspase-3 and terminal deoxynucleotidyl transferase (TdT) assay (TUNEL) for detection of internucleosomal DNA fragmentation.
Microscopic evaluation to assess the presence and cellular co-localization of active caspase-3 and TUNEL-positive cells.
In human ovarian tissue, no apoptosis was observed in primordial and primary follicles. Apoptosis in granulosa cells was detected only in atretic antral follicles. Granulosa cells classified as apoptotic on the basis of their morphologic features contained a single condensed nucleus, multiple nuclear fragments, or apoptotic bodies. All apoptotic granulosa cells expressed active caspase-3, but only few contained fragmented DNA detected with the TUNEL method. The expression of active caspase-3 was also demonstrated in human granulosa cells of preovulatory follicles obtained from patients undergoing IVF.
Caspase-3 dependent apoptosis occurs in human granulosa cells and activates when follicles begin to leave the resting pool. After initial formation of the antrum, activation of caspase-3 is a normal physiologic process of the follicle during atresia and luteinization. Higher numbers of granulosa cells positive with caspase-3 than cells positive with TUNEL suggest an earlier activation of caspase-3 compared with the DNA fragmentation detected by TUNEL assay and also a longer detection period of caspase-3 than DNA fragmentation in apoptotic granulosa cells.
Bladder cancer is the fourth most commonly diagnosed malignancy in men worldwide and has one of the highest recurrence rates of all cancers. This cancer type is unique because chronic inflammation ...caused by Schistosoma haematobium can cause bladder cancer, while inflammation induced by Bacillus Calmette Guerin is the therapeutic cornerstone for this cancer type. Activation of proinflammatory IL-6/Stat3 axis promotes the development of different cancers by acting on cancer cells as well as by modulating cancer microenvironment. Using a genetic and pharmacological approach in a mouse model, we demonstrated the importance of IL-6 and Stat3 signaling in bladder cancer. Our findings show that pharmacological inhibition of Stat3 with WP1066 effectively delays progression and invasiveness of bladder cancer in N-butyl-N-(4-hydroxybutyl) nitrosamine-induced mouse model. Moreover, either IL-6 blockade or Stat3 inhibition sensitized bladder cancer to anti-PD-L1 immune therapy. Taken together, our study demonstrates an important role of IL-6/Stat3 signaling in bladder cancer and creates a rationale for testing the therapeutic potential of Stat3 inhibitors in human MIBC both alone or in combination with anti-PD-L1 and anti-IL-6 therapy.
•Stat3 is activated during bladder carcinogenesis in the BBN mouse model.•Genetic ablation and systemic inhibition of Stat3 impair bladder carcinogenesis.•Ablation of IL-6 promotes BC through an aberrant immune response.•Targeting IL-6/Stat3 and immune checkpoint inhibitors prevents BC invasiveness.•Stat3 inhibition has therapeutic potential in human MIBC.
Abstract Background The aim of this study was to compare lateral thermal damage of mesoappendix and appendiceal base using three different instruments for sealing and cutting of mesoappendix. ...Materials and methods A total number of 99 patients (54 males and 45 females) who underwent laparoscopic appendectomy because of suspected appendicitis between December 2013 and May 2015 were enrolled in the study. The patients were divided in three groups based on instrument used for sealing of mesoappendix: group 1 (Ultracision; n = 36), group 2 (LigaSure; n = 32), and group 3 (MiSeal; n = 31). Lateral thermal damage, intraoperative and postoperative complications, duration of surgery, hospital stay, and economic value were compared within groups. Results The median age of patients was 14 y (range 3-17). A histopathologic analysis revealed a positive diagnosis of appendicitis in 84 patients (85%). The median lateral thermal damage on appendiceal base using Ultracision, LigaSure, and MiSeal was 0.10 mm, 0.16 mm, and 0.10 mm respectively, and on mesoappendix, 0.08 mm, 0.13 mm, and 0.08 mm, respectively. Significantly higher thermal damage was found on mesoappendix ( P = 0.015) and appendiceal base ( P = 0.012) in patients treated with LigaSure than in patients from other groups. There were no statistical differences among the groups regarding intraoperative and postoperative complications ( P = 0.098). No significant difference in thermal damage between appendicitis and nonappendicitis group was found ( P = 0.266). Conclusions Using of Ultracision, LigaSure, and MiSeal for sealing of mesoappendix in laparoscopic appendectomy in children is safe and useful. LigaSure produces significantly greater lateral thermal damage compared with other instruments.
Urinary bladder cancer (BC) inflicts a significant impairment of life quality and poses a high mortality risk. Schistosoma haematobium infection can cause BC, and the urinary microbiota of BC ...patients differs from healthy controls. Importantly, intravesical instillation of the bacterium Bacillus Calmette-Guerin stands as the foremost therapy for non-muscle invasive BC. Hence, studying the receptors and signaling molecules orchestrating bacterial recognition and the cellular response in the context of BC is of paramount importance. Thus, we challenged Toll-like receptor 4 (Tlr4) and myeloid differentiation factor 88 (Myd88) knock-out (KO) mice with N-butyl-N-(4-hydroxylbutyl)-nitrosamine (BBN), a well-known urinary bladder carcinogen. Gut microbiota, gene expression, and urinary bladder pathology were followed. Acute exposure to BBN did not reveal a difference in bladder pathology despite differences in the animal’s ability to recognize and react to bacteria. However, chronic treatment resulted in reduced cancer invasiveness among Myd88KO mice while the absence of functional Tlr4 did not influence BC development or progression. These differences correlate with a heightened abundance of the Faecalibaculum genus and the lowest microbial diversity observed among Myd88KO mice. The presented data underscore the important role of microbiota composition and MyD88-mediated signaling during bladder carcinogenesis.
Different pathophysiological models provide insight into the important role of CD83
dendritic cells (DCs) in the pathogenesis of Crohn's disease (CD) and ulcerative colitis (UC). There were 154 ...subjects included in this study: 60 with UC, 19 with CD and 75 in the control group. Colonic biopsy was performed in all subjects. Specimens were incubated with a primary anti-CD83 antibody. Intraepithelial DCs per 100 enterocytes were counted. The results were analysed according to demographic data, type of IBD and histological inflammation pattern. The odds ratio for CD83
DCs=0 in the UC group was 3.4 times higher than that in the control group (OR = 3.4; 95% CI: 1.63-7.14; p = 0.001), and the odds ratio for CD83
DCs ≥1 in the CD group was 5.3 times higher than that in the UC group (OR = 5.3; 95% CI: 1.4-20.2; p = 0.014). The odds ratio for CD83
DCs=0 in the acute inflammation group was 2.7 times higher than that in the group without inflammation (OR = 2.7; 95% CI: 1.2-5.9; p = 0.011). In the group of patients with CD and acute inflammation (n = 11), there was only one subject without CD83
DCs (p = 0,024). These results suggest an association of CD83
DCs with the type of IBD and the histological inflammation pattern.
Colorectal cancer is the second most common cause of cancer-related mortality in adults. Understanding colorectal tumorigenesis at both the cellular and molecular levels is crucial for developing ...effective treatment options. Forty-one biopsy samples from patients with metastatic CRC (mCRC) were collected at Split University Hospital in Croatia. A total of 41 patients (21 with microsatellite unstable tumours and 20 with microsatellite stable tumours) were randomly included in the study. Immunolabelling of cGAS and STING in metastatic CRC was performed and further complemented by histological classification, tumour grade, and KRAS, NRAS, and BRAF mutational status of mCRC. In bivariate analysis, elevated expression of cGAS and STING was positively associated with MSI-H colon cancer (Fisher's exact test, both
= 0.0203). Combined expression analysis of cGAS and STING showed a significantly higher percentage of patients with mCRC MSI-H with a fully or partially activated cGAS-STING signalling pathway (chi-square test,
= 0.0050). After adjusting for age, sex, and STING expression, increased cGAS expression remained significantly associated with MSI-H colon cancer in a multiple logistic regression model (β = 1.588, SE = ±0.799,
= 0.047). The cGAS-STING signalling axis represents a compelling new target for optimization of immune checkpoint inhibitor therapeutic approaches in patients with MSI-H stage IV CRC.
Laryngeal cancer is the second most common malignancy of the head and neck, worldwide. Immunotherapy targeting checkpoint inhibitors has been approved for the treatment of patients with recurrent or ...metastatic laryngeal cancer but has a relatively low response rate and outcomes that leave many patients underserved. Targeting the cGAS-STING signaling pathway can potentially improve the activation of immune effector cells, although its role in the development and progression of laryngeal cancer has not yet been investigated in depth. Fifty-nine tumor samples from patients with pathologically confirmed squamous cell carcinoma of the larynx, stage I-IV non-metastatic disease, who were treated at the University Hospital of Split, were immunohistochemically stained for the expression of STING, cGAS, CD8, CD68, and CD163. Elevated tumor cell-intrinsic STING expression was positively associated with stage IV (
= 0.0031), pT3, and pT4 laryngeal cancers (
= 0.0336) as well as with higher histological grades (G2 and G3) (
= 0.0204) and lymph node-positive tumors (
= 0.0371). After adjusting for age, sex, location, and cGAS expression, elevated STING expression was significantly associated with stage IV cancer in a multiple logistic regression model (β = 1.849, SE = ±0.8643,
= 0.0324). Elevated STING expression represents a potentially favorable predictive biomarker for new therapeutic approaches involving STING agonists combined with immunotherapy and DNA-damaging agents (radiotherapy, cisplatin, and PARP inhibitors) in laryngeal cancer.
The development of colorectal cancer is known to be characterized by a sequence of events during which normal colonic epithelium gradually transforms to carcinoma, the adenoma-carcinoma sequence. ...Apoptosis plays an important role in the development and maintenance of tissue homeostasis. Currently, there is no agreement in the literature about the prognosis of apoptosis in colorectal cancer. The number of studies examining the expression of caspases in colorectal cancer is very limited, and they have not examined any correlation between expression and patient survival. This study included histologic samples from 179 patients diagnosed with colon cancer. We used the TdT-mediated X-dUTP nick end labeling method and caspase-3 labeling to identify the degree of apoptosis. Our results show that lower apoptotic index measured by TdT-mediated X-dUTP nick end labeling method and lower immnuhistochemical expression of caspase-3 is associated with shorter disease-free survival and overall survival. However, only apoptotic index is proven to be an independent survival indicator. The results of our study are consistent with the proposed models of carcinogenesis of colorectal cancer that emphasize resistance to apoptosis as a decisive factor in the progression of the disease and resistance to treatment.
Bladder cancer (BC) is the ninth leading cause of cancer death with one of the highest recurrence rates among all cancers. One of the main risks for BC development is exposure to nitrosamines present ...in tobacco smoke or in other products. Aberrant epigenetic (DNA methylation) changes accompanied by deregulated gene expression are an important element of cancer pathogenesis. Therefore, we aimed to determine DNA methylation signatures and their impacts on gene expression in mice treated with N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN), a carcinogen similar to compounds found in tobacco smoke. Following BBN administration mice developed non-invasive or invasive bladder cancers. Surprisingly, muscle- and neuronal-related pathways emerged as the most affected in those tumors. Hypo- and hypermethylation changes were present within non-invasive BC, across CpGs mapping to the genes involved in muscle- and neuronal-related pathways, however, methylation differences were not sufficient to affect the expression of the majority of associated genes. Conversely, invasive tumors displayed hypermethylation changes that were linked with alterations in gene expression profiles. Together, these findings indicate that bladder cancer progression could be revealed through methylation profiling at the pre-invasive cancer stage that could assist monitoring of cancer patients and guide novel therapeutic approaches.
Purpose
The aim of this study was to investigate the effect of urapidil and low-molecular weight heparin (LMWH) on testicular torsion–detorsion injury in rats.
Methods
Thirty-two male Sprague–Dawley ...rats were used. In the torsion–detorsion (T/D) group, the left testis was twisted at 720° for 3 h. After 3 h of reperfusion, the testis was removed. Urapidil or LMWH was administered intraperitoneally 30 min before detorsion in the treatment groups.
Results
Unilateral testicular torsion–detorsion caused significant increases in the malondialdehyde level and apoptosis and significant decreases in the superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities in ipsilateral testes (
p
< 0.001). The rats treated with urapidil had a significant decrease in the malondialdehyde level and apoptosis and significant increases in the SOD and GPx activities in ipsilateral testes compared to the T/D group (
p
< 0.001). Animals treated with LMWH showed non-significant reductions in malondialdehyde levels and apoptosis compared to the T/D group. In addition, no significant difference in the SOD activities (
p
= 0.52) between the groups was found. The increase in the GPx activities was significant in the LMWH group compared to the T/D group (
p
< 0.001).
Conclusion
The administration of urapidil before detorsion prevents ischemia/reperfusion cellular damage in testicular tissue. LMWH was not found to have a beneficial effect on testicular T/D injury in rats.
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