Translocator Protein Modulation by 4'-Chlorodiazepam and NO Synthase Inhibition Affect Cardiac Oxidative Stress, Cardiometabolic and Inflammatory Markers in Isoprenaline-Induced Rat Myocardial Infarction.

The possible cardioprotective effects of translocator protein (TSPO) modulation with its ligand 4'-Chlorodiazepam (4'-ClDzp) in isoprenaline (ISO)-induced rat myocardial infarction (MI) were evaluated, alone or in the presence of L-NAME. Wistar albino male rats (b.w. 200-250 g, age 6-8 weeks) were divided into 4 groups (10 per group, total number N = 40), and certain substances were applied: 1. ISO 85 mg/kg b.w. (twice), 2. ISO 85 mg/kg b.w. (twice) + L-NAME 50 mg/kg b.w., 3. ISO 85 mg/kg b.w. (twice) + 4'-ClDzp 0.5 mg/kg b.w., 4. ISO 85 mg/kg b.w. (twice) + 4'-ClDzp 0.5 mg/kg b.w. + L-NAME 50 mg/kg b.w. Blood and cardiac tissue were sampled for myocardial injury and other biochemical markers, cardiac oxidative stress, and for histopathological evaluation. The reduction of serum levels of high-sensitive cardiac troponin T hs cTnT and tumor necrosis factor alpha (TNF-α), then significantly decreased levels of serum homocysteine Hcy, urea, and creatinine, and decreased levels of myocardial injury enzymes activities superoxide dismutase (SOD) and glutathione peroxidase (GPx) as well as lower grades of cardiac ischemic changes were demonstrated in ISO-induced MI treated with 4'-ClDzp. It has been detected that co-treatment with 4'-ClDzp + L-NAME changed the number of registered parameters in comparison to 4'-ClDzp group, indicating that NO (nitric oxide) should be important in the effects of 4'-ClDzp.

Immunohistochemical expression of caspases 9 and 3 in adenoid cystic carcinoma of salivary glands and association with clinicopathological parameters.

PURPOSE: Adenoid cystic carcinoma (ACC) is one of the most common malignant salivary gland tumors. It is characterized by a high rate of recurrence, perineural invasion and development of distant metastases many years after removal of the primary tumor. Disorders of the induction of apoptosis and its cascade reactions where caspases are involved may be significant in the pathogenesis of this tumor. METHODS: The immunohistochemical expression of caspase 9 and caspase 3 was analyzed by tissue microarray (TMA) in 50 cases of ACC in relation with different clinicopathological parameters (gender, age, localization, histological type and overall survival). RESULTS: Caspase 9 was expressed in the cytoplasm and nuclei of ACC tumor cells with varying degrees of staining intensity (1+, 6%; 2+, 54%, 3+, 40%). Comparison of caspase 9 expression in tumor cells with clinicopathological parameters (gender, age, localization, histological type and overall survival) showed no statistically significant difference except that the expression was more pronounced in females. Caspase 3 was expressed in the cytoplasm of tumor cells with varying degrees of staining intensity (1+, 22%; 2+, 36%; 3+, 42%). No correlation between the expression of caspase 3 and clinicopathological parameters was noticed. CONCLUSIONS: The expression of caspases 9 and 3 in ACC of the salivary glands can contribute in the better characterization of molecules involved in apoptosis of tumor cells.

BCL2 positive and BCL6 negative diffuse large B cell lymphoma patients benefit from R-CHOP therapy irrespective of germinal and non germinal center B cell like subtypes.

PURPOSE: Despite major advances in the treatment of diffuse large B cell lymphoma (DLBCL), approximately one third of the patients progress or die, suggesting the existence of additional oncogenic events. The purpose of this study was to evaluate the prognostic value of the "Hans classifier", and BCL2 and MYC protein expression and gene alterations in DLBCL patients treated with CHOP or R-CHOP chemotherapy over a 5-year period. Furthermore, we tried to correlate these parameters with the International Prognostic Index (IPI). METHODS: The immunohistochemical (IHC) expression of CD10, BCL6, MUM1 and BCL2 on paraffin-embedded formalin-fixed tumor samples from 103 centroblastic DLBCLs was analyzed. IHC expression of MYC and fluorescence in situ hybridization (FISH) for MYC and BCL2 gene alterations was performed on 67 samples using the tissue microarray (TMA) method. RESULTS: The Hans algorithm was not predictive of survival in both therapy groups. No significant difference in BCL2 and MYC alterations or MYC protein expression in relation to complete response (CR), event-free survival (EFS) and overall survival (OS) was observed in our study. High IPI correlated significantly with poor outcome and it was identified as independent prognostic factor for OS and EFS (both p=0.000). The 5-year OS was 61% in the R-CHOP compared to 38% in the CHOP group (p=0.007). Rituximab significantly improved the OS in the BCL2 positive (60 vs 29%, p=0.008), and the BCL6 negative (73 vs 25%, p=0.001) cases. CONCLUSION: IPI is an independent prognosticator for DL-BCL patients and the addition of rituximab significantly improved survival. Furthermore, patients with BCL2+ and BCL6-DLBCL benefited from R-CHOP.

Case Report: Giant cholesterol granuloma in the anterior mediastinum.

Cholesterol granuloma is a rare entity, which can develop in many regions of the body, accounting at most 1% of all mediastinal tumors. Etiology of this granuloma is still not clearly understood. The gold standard choice of treatment for cholesterol granuloma is total surgical resection. Symptomatic mediastinum granuloma can be easily diagnosed, but if mass effect is not evident then diagnosis of this tumor is really challenging. We present a rare case of huge cholesterol granuloma in the anterior mediastinum of the patient who underwent on elective coronary artery graft bypass surgery.

Derivation of benchmark doses for male reproductive toxicity in a subacute low-level Pb exposure model in rats.

The objectives of the study were to simulate low-level Pb exposure scenario in an animal model and to examine reproductive adverse effects. Based on obtained data, we have performed Benchmark dose (BMD)-response modelling. Male Wistar rats were randomized in seven groups (n = 6): one control and six treated with: 0.1, 0.5, 1, 3, 7, and 15 mg Pb/kg body weight, daily for 28 days by oral gavage. The rats were sacrificed and the blood and testes were used for further analysis of testosterone levels in serum, testicular essential metal levels and histological analysis. The Pb treatment led to a dose-dependent decrease of serum testosterone levels with a negative trend (BMDI 0.17-6.13 mg Pb/kg). Increase of Zn (dose-dependent, BMDI 0.004-19.7 mg Pb/kg) and Cu and a decrease of Mn testicular levels were also detected with unscathed histology of the testes. The presented results might be used in further evaluation of the point of departure in human health risk assessment for Pb.

Syringocystadenoma Papilliferum in the External Ear Canal.

Out of all benign tumors of the ceruminous glands, syringocystadenoma papilliferum is the rarest and represents only 2% of cases. It is an extremely rare benign tumor that originates from modified apocrine sweat glands. The aim of this paper was to present, according to our findings, the 18th case of syringocystadenoma papilliferum in the external auditory canal, with a detailed review of its clinical, radiological and histomorphological characteristics. A 59-year-old man reported to our clinic due to a 5×5 mm papillomatous growth at the entrance to the right external auditory canal. Histopathology indicated, after an excisional biopsy, that it was a syringocystadenoma papilliferum. The resection lines were free of tumor tissue, and the patient has no signs of tumor recurrence. Although rare, it should be considered as a differential diagnosis of lesions in this region. Complete excision is mandatory in order to avoid recurrence and potential malignant alteration.

Utility of CK8, CK10, CK13, and CK17 in Differential Diagnostics of Benign Lesions, Laryngeal Dysplasia, and Laryngeal Squamous Cell Carcinoma.

There are no reliable immunohistochemical markers for diagnosing laryngeal squamous cell carcinoma (SCC) or diagnosing and grading laryngeal dysplasia. We aimed to evaluate the diagnostic utility of CK8, CK10, CK13, and CK17 in benign laryngeal lesions, laryngeal dysplasia, and laryngeal SCC. This retrospective study included 151 patients diagnosed with laryngeal papilloma, laryngeal polyps, laryngeal dysplasia, and laryngeal SCC who underwent surgical treatment between 2010 and 2020. Immunohistochemistry (IHC) was carried out using specific monoclonal antibodies against CK8, CK10, CK13, and CK17. Two experienced pathologists performed semi-quantitative scoring of IHC positivity. The diagnostic significance of the markers was analyzed. CK13 showed a sensitivity of 100% and a specificity of 82.5% for distinguishing between laryngeal SCC and laryngeal dysplasia and benign lesions. CK17 showed a sensitivity of 78.3% and specificity of 57.1% for the detection of laryngeal SCC vs. laryngeal dysplasia. CK10 showed a sensitivity of 80.0% for discriminating between low-grade and high-grade dysplasia, and a specificity of 61.1%. Loss of CK13 expression is a reliable diagnostic tool for diagnosing laryngeal lesions with malignant potential and determining resection lines. In lesions with diminished CK13 expression, CK17 could be used as an auxiliary immunohistochemical marker in diagnosing laryngeal SCC. In CK13-negative and CK17-positive lesions, CK10 positivity could be used to determine low-grade dysplasia. CK8 is not a useful IHC marker in differentiating between benign laryngeal lesions, laryngeal dysplasia, and laryngeal SCC.

Comprehensive insight into the neurotoxic mechanisms of low dose Pb exposure in Wistar rats: Benchmark dose analysis.

Lead (Pb) is a toxic metal that affects almost all human's system and organs, with the nervous system as the most sensitive. Better understanding of the Pb neurobehavioral effects and neurotoxicity requires realistic study scenarios based on low level exposure. The aim of this study was to determine neurotoxic effects and mechanisms of Pb in six low doses and to establish dose-response relationship for these effects and related Benchmark dose (BMD). Forty-two, male albino Wistar rats were randomized into seven groups, control and Pb-exposed: 0.1, 0.5, 1, 3, 7 and 15 mg Pb/kg body weight/day (oral gavage) for 28 days. Behavioural tests (Elevated plus maze test, Spontaneous locomotor activity test and Novel object recognition test) were conducted in the last week of experiment, in the control, lower (0.5 mg Pb/kg), middle (3 mg Pb/kg) and higher (15 mg Pb/kg) dose groups. The acetylcholinesterase activity, oxidative status and essential elements levels (Cu, Zn, Mn and Fe) were measured in brain tissue along with histological analyses. External and internal dose-response analyses were performed using PROASTweb 70.1 software. The results have shown that subacute exposure to very low doses of Pb resulted in memory deficits in rats that was accompanied with acetylcholinesterase enzyme activity decrease. The observed hyperactive behaviour was accompanied by dose-dependent induction of brain oxidative stress and Zn elevation. The histological alterations in Purkinje cells were only detected in the group treated with the highest Pb dose. The lowest BMD considering entire oxidative status was calculated based on total oxidative status (4.5e-06 mg Pb/kg b.w./day). The findings reported in our study may be beneficial in further evaluating the health consequences and human health risk assessment of low-level Pb exposure.

Environmental cadmium exposure and pancreatic cancer: Evidence from case control, animal and in vitro studies.

Although profoundly studied, etiology of pancreatic cancer (PC) is still rather scarce. Some of established risk factors of PC are connected to an increased cadmium (Cd) body burden. Hence, the aim of this study was to investigate the role of this environmental pollutant in PC development by conducting human observational, experimental and in vitro studies. The case-control study included 31 patients with a histologically based diagnosis of exocrine PC subjected to radical surgical intervention as cases and 29 accidental fatalities or subjects who died of a nonmalignant illness as controls. Animal study included two treated groups of Wistar rats (15 and 30 mg Cd/kg b.w) and untreated control group, sacrificed 24 h after single oral exposure. In in vitro study pancreas hTERT-HPNE and AsPC-1 cells were exposed to different Cd concentrations corresponding to levels measured in human cancerous pancreatic tissue. Cd content in cancer tissue significantly differed from the content in healthy controls. Odds ratio levels for PC development were 2.79 (95% CI 0.91-8.50) and 3.44 (95% CI 1.19-9.95) in the third and fourth quartiles of Cd distribution, respectively. Animal study confirmed Cd deposition in pancreatic tissue. In vitro studies revealed that Cd produces disturbances in intrinsic pathway of apoptotic activity and the elevation in oxidative stress in pancreatic cells. This study presents three different lines of evidence pointing towards Cd as an agent responsible for the development of PC.

Toxic Effect of Acute Cadmium and Lead Exposure in Rat Blood, Liver, and Kidney.

Background: Cadmium and lead are widespread and non-biodegradable pollutants of great concern to human health. In real life scenarios, we are exposed to mixtures of chemicals rather than single chemicals, and it is therefore of paramount importance to assess their toxicity. In this study, we investigated the toxicity of Cd and Pb alone and as a mixture in an animal model of acute exposure. Methods: Experimental groups received a single treatment of aqueous solution of Cd-chloride (15 and 30 mg/kg body weight (b.w.) and Pb-acetate (150 mg/kg b.w.), while the mixture group received 15 mg Cd/kg b.w. and 150 mg Pb/kg b.w. Toxic effects of individual metals and their mixture were investigated on hematological and biochemical parameters, and the redox status in the plasma, liver, and kidneys of treated Wistar rats. Results: Tissue-specific changes were recorded in various parameters of oxidative damage, while the accumulation of metals in tissues accompanied the disturbances of both hematological and biochemical parameters. It was observed that the level of toxic metals in tissues had a different distribution pattern after mixture and single exposure. Conclusions: Comprehensive observations suggest that exposure to Cd and Pb mixtures produces more pronounced effects compared to the response observed after exposure to single metal solutions. However, further research is needed to confirm toxicokinetic or toxicodynamic interactions between these two toxic metals in the organisms.

Expression of Minichromosome Maintenance Proteins in Actinic Keratosis and Squamous Cell Carcinoma.

Minichromosome maintenance (MCM) proteins are a group of proteins involved in DNA replication and cell-cycle regulation. Because they are associated with DNA through G1 into S phase, MCM proteins are potentially specific indicators of cell proliferation that could be valuable markers of dysplasia, and preinvasive and invasive malignant tumors. To analyze MCM protein expression patterns in actinic keratosis (AK), Bowen disease (BD), and cutaneous squamous cell carcinoma (SCC), we performed immunohistochemical staining of MCM2, -5, and -7 on tissue microarray blocks from 91 AK, 50 BD, and 174 SCC samples. The distribution and semiquantitatively assessed number of positive cells were analyzed in relation to the type of the lesion and the SCC prognostic parameters (grade, diameter, and thickness). Basal expression of all 3 proteins was observed more frequently in AK, whereas the distribution in BD was predominantly diffuse (P<0.001). All 3 proteins showed peripheral distribution in most well-differentiated SCC and diffuse distribution in poorly differentiated tumors (P<0.001). Using the 50% cut-off value, there was a statistically significant difference among AK, BD, and SCC (P<0.001). In addition, all MCM proteins showed highly significant differences (P<0.001) between well-differentiated SCC and both moderately and poorly differentiated SCC. The diffuse distribution and 50% cut-off value of positive cells revealed statistically significant associations of all MCM proteins with SCC thicker than 6 mm. Our results suggest a role for MCM proteins in the progression of in situ keratinocytic lesions and their association with high-risk features in SCC.

The Role of Lymphocyte to Monocyte Ratio, Microvessel Density and HiGH CD44 Tumor Cell Expression in Non Hodgkin Lymphomas.

Prognostic significance of immune microenvironment has been emphasized using the most advanced analysis, with consecutive attempts to reveal prognostic impact of this findings. The aim of this study was to compare and define prognostic significance of clinical parameters, microvessel density (MVD) in tumour tissue and expression of CD44s as adhesive molecule on tumour cells in diffuse large B cell lymphoma-DLBCL, primary central nervous system DLBCL-CNS DLBCL and follicular lymphoma-FL. A total of 202 histopathological samples (115 DLBCL/65 FL/22 CNS DLBCL) were evaluated. Overall response (complete/partial remission) was achieved in 81.3 % DLBCL patients, 81.8 % primary CNS DLBCL and 92.3 % FL. Absolute lymphocyte count-ALC/Absolute monocyte count-AMC >2.6 in DLBCL and ALC/AMC ≥ 4.7 in FL were associated with better event-free survival (EFS) and overall survival (OS) (p < 0.05). In DLBCL, MVD > 42 blood vessels/0.36 mm(2) correlated with primary resistant disease (p < 0.0001), poorer EFS and OS (p = 0.014). High CD44s expression in FL correlated with inferior EFS and OS (p < 0.01). In DLBCL, multivariate Cox regression analysis showed that ALC/AMC was independent parameter that affected OS (HR 3.27, 95 % CI 1.51-7.09, p = 0.003) along with the NCCN-IPI (HR 1.39, 95 % CI 1.08-1.79, p = 0.01). Furthermore, in FL, ALC/AMC mostly influenced OS (HR 5.21, 95 % CI 1.17-23.21, p = 0.03), followed with the FLIPI (HR 3.98, 95 % CI 1.06-14.95, p = 0.041). In DLBCL and FL, ALC/AMC is simple and robust tool that is, with current prognostic scores, able to define long-term survival and identify patients with inferior outcome. The introduction of immunochemotherapy might altered the prognostic significance of microenvionmental biomarkers (MVD and CD44s).

Oncogene-Induced Senescence in Pituitary Adenomas--an Immunohistochemical Study.

Oncogene-induced senescence (OIS) serves as an initial barrier to cancer development, being proposed as a possible explanation for the usually benign behavior of the pituitary adenomas. We aimed to explore the immunohistochemical expression of the OIS markers, senescence-associated lysosomal ß-galactosidase (SA-ß-GAL), p16, and p21 in different types of 345 pituitary adenomas and compared it with the expression in the normal pituitary and in the specimens from the repeated surgeries. SA-ß-GAL was overexpressed in the pituitary adenomas, compared to the normal pituitaries. Growth hormone (GH) producing adenomas showed the strongest SA-ß-GAL, with densely granulated (DG)-GH adenomas more reactive than the sparsely granulated (SG). Nuclear p21 was decreased in the adenomas, except for the SG-GH adenomas that had higher p21 than the normal pituitaries and the other adenomas. p16 was significantly lower in the adenomas, without type-related differences. SA-ß-GAL was slightly lower and p16 slightly higher in the recurrences. Our findings indicate alterations of the senescence program in the different types of pituitary adenomas. Activation of senescence in the pituitary adenomas presents one possible explanation for their usually benign behavior, at least in the GH adenomas that show a synchronous increase of two OIS markers. However, subdivision into GH adenoma subtypes reveals differences that reflect complex regulatory mechanisms influenced by the interplay between the granularity pattern and the hormonal factors, with possible impact on the different clinical behavior of the SG- and DG-GH adenoma subtypes. p16 seems to have a more prominent role in the pituitary tumorigenesis than in the senescence. Recurrent growth in a subset of the pituitary adenomas is not associated with consistent changes in the senescence pattern.

Immunohistochemical Expression of p16 and p21 in Pituitary Tissue Adjacent to Pituitary Adenoma versus Pituitary Tissue Obtained at Autopsy: Is There a Difference?

Normal pituitary tissue is frequently used for comparison with protein expression in tumor tissue, being obtained either at surgery or at autopsy. p16 and p21 proteins are cyclin-dependent kinase inhibitors, belonging to INK4 and Cip/Kip family, respectively. Their expression is increased in response to DNA damage or other cellular stressors, resulting in the activation of cell cycle checkpoints. They also play important roles in cellular senescence. The purpose of this study was to investigate differences in p16 and p21 immunohistochemical expression in normal pituitary tissue adjacent to pituitary adenoma obtained during neurosurgical procedure with pituitary tissue obtained at autopsy, from patients who died from non-endocrinological diseases. Our results show significant difference in p16 nuclear and p21 cytoplasmic immunohistochemical expression between two types of normal pituitary tissues. One of the reasons for this difference could be the age of subjects because those who underwent autopsy for a non-endocrinological disease were significantly older than subjects who underwent neurosurgery for a pituitary adenoma. Our finding that differences are probably not influenced by postmortem changes is supported by no significant correlation between postmortem interval and immunohistochemical p16 and p21 expression. The influence of the presence of a pituitary adenoma could not be evaluated in these specimens.