BCL-2 and γ-H2AX immunostaining profile in urothelial bladder cancer prognosis.

Urothelial bladder carcinoma (UBC) is a malignant neoplasm of the urinary tract that is highly prevalent worldwide and has a high rate of tumor recurrence. It is known that the BCL2 apoptosis regulator (BCL-2) gene encodes a mitochondrial protein that regulates programmed death cells by apoptosis. In contrast, the H2A.X histone variant (H2AX) gene encodes a histone responsible for regulating and signaling genomic instability processes. The present study aimed to analyze the immunostaining profiles of BCL-2 and γ-H2AX proteins in tissue samples (n=80) from UBC patients (muscle-invasive MI; and non-muscle invasive NMI) using indirect immunohistochemistry and to correlate the results with prognostic and clinical parameters. BCL-2 protein expression was cytoplasmic and absent in half of the samples, including the MI and NMI groups. Strong nuclear expression was observed for γ-H2AX, predominant in the MI samples. The immunostaining profile of both proteins was not associated with tumor recurrence or invasion, and no significant associations were found in relation to prognosis (tumor grade, pathological staging). No significant correlation was found between protein profiles in malignant tissue. All in all, BCL-2 and γ-H2AX did not prove to be candidate markers for UBC clinical management in the present sample, despite their expression in malignant bladder tissue.

Descriptive study of perineuronal net in enteric nervous system of humans and mice.

Perineuronal nets (PNN) are highly specialized structures of the extracellular matrix around specific groups of neurons in the central nervous system (CNS). They play functions related to optimizing physiological processes and protection neurons against harmful stimuli. Traditionally, their existence was only described in the CNS. However, there was no description of the presence and composition of PNN in the enteric nervous system (ENS) until now. Thus, our aim was to demonstrate the presence and characterize the components of the PNN in the enteric nervous system. Samples of intestinal tissue from mice and humans were analyzed by RT-PCR and immunofluorescence assays. We used a marker (Wisteria floribunda agglutinin) considered as standard for detecting the presence of PNN in the CNS and antibodies for labeling members of the four main PNN-related protein families in the CNS. Our results demonstrated the presence of components of PNN in the ENS of both species; however its molecular composition is species-specific.

Deposition of collagen III and alterations in basement membrane integrity as candidate prognostic markers in prostate cancer.

The extracellular matrix surrounding the tumor undergoes changes in its organization during the metastasis process. The present study aims to quantify total collagen, collagen I (Col I) and collagen III (Col III), analyze the alignment of collagen fibers and assess the basement membrane integrity in samples from patients with metastatic and non-metastatic prostate cancer. Tissue samples from 60 patients were classified into groups based on prognostic parameters: better prognosis (n = 20), worse prognosis without metastasis (n = 23) and metastatic (n = 17). Picrosirius red with further analysis under polarizing microscope was used to quantify (with validation using immunohistochemistry) and analyze collagen alignment, and Periodic Acid Schiff staining was used to analyze the basement membrane integrity. The Col I/Col III ratio was found to be higher in the metastatic group than in the groups with better prognosis (p = 0.012) and worse prognosis without metastasis (p = 0.018). Basement membrane integrity constitution in malignant tumor tissue differed from that of adjacent non-tumor tissue (p < 0.001). Moreover, the worsening in the tumor tissue integrity was positively correlated with worse prognostic parameters. All in all, absence of Col III and basement membrane integrity might be indicators of poor prognosis in prostate cancer.

Perineuronal net in the extrinsic innervation of the distal colon of mice and its remodeling in ulcerative colitis.

The perineuronal net (PNN) is a well-described highly specialized extracellular matrix structure found in the central nervous system. Thus far, no reports of its presence or connection to pathological processes have been described in the peripheral nervous system. Our study demonstrates the presence of a PNN in the spinal afferent innervation of the distal colon of mice and characterizes structural and morphological alterations induced in an ulcerative colitis (UC) model. C57Bl/6 mice were given 3% dextran sulfate sodium (DSS) to induce acute or chronic UC. L6/S1 dorsal root ganglia (DRG) were collected. PNNs were labeled using fluorescein-conjugated Wisteria Floribunda (WFA) l lectin, and calcitonin gene-related peptide (CGRP) immunofluorescence was used to detect DRG neurons. Most DRG cell bodies and their extensions toward peripheral nerves were found surrounded by the PNN-like structure (WFA+), labeling neurons' cytoplasm and the pericellular surfaces. The amount of WFA+ neuronal cell bodies was increased in both acute and chronic UC, and the PNN-like structure around cell bodies was thicker in UC groups. In conclusion, a PNN-like structure around DRG neuronal cell bodies was described and found modulated by UC, as changes in quantity, morphology, and expression profile of the PNN were detected, suggesting a potential role in sensory neuron peripheral sensitization, possibly modulating the pain profile of ulcerative colitis.

Metalloproteinase 9 immunostaining profile is positively correlated with tumor grade, extraprostatic extension and biochemical recurrence in prostate cancer.

Metastasis is the main problem in the treatment of prostate cancer (PCa), and for it to occur, proteolytic enzymes must remodel the extracellular matrix (ECM) surrounding the tumor. The most important group of enzymes with this action include the matrix metalloproteinases (MMPs), which act on various substrates cleaving ECM components. The present study aimed to evaluate the protein immunostaining profiles of matrix metalloproteinase 2 (MMP-2) and 9 (MMP-9) in PCa Brazilian patients using the indirect immunohistochemical methodology. The tissue samples (n = 178), 60 from malignant tumor, 58 from adjacent non-tumor, and 60 from ECM, were evaluated according to the immunostaining intensity. The malignant tumor cytoplasmic MMP-2 immunostaining was more intense than in ECM (p = 0.001), but it did not correlate with any clinical-pathological parameter. The MMP-9 staining was similar in tumor cytoplasm, adjacent non-tumor cytoplasm and ECM, but showed significant positive correlations with ISUP grade (p = 0.044; Tau=0.249), extraprostatic extension (p = 0.025; Tau=0.309), and biochemical recurrence (p = 0.048; Tau=0.306). A significant positive correlation was also observed between MMP-2 and MMP-9 in all cell compartments analyzed. Although further research is warranted to elucidate the precise mechanisms underlying these observations, our findings suggest MMP-9 as a promising candidate marker for tissue invasion that could be used in predicting the progression and prognosis of PCa.

Tissue immunostaining of candidate prognostic proteins in metastatic and non-metastatic prostate cancer.

PURPOSE: Prostate cancer (PCa) lacks specific markers capable of distinguishing aggressive tumors from those with indolent behavior. Therefore, the aim of this study was to evaluate the immunostaining of candidate proteins (PTEN, AKT, TRPM8, and NKX3.1) through the immunohistochemistry technique (IHC) on patients with metastatic and non-metastatic PCa. METHODS: Tissues from 60 patients were divided into three groups categorized according to prognostic parameters: better prognosis (n = 20), worse prognosis (n = 23), and metastatic (n = 17). Immunostaining was analyzed by a pathologist and staining classifications were considered according to signal intensity: (0) no staining, (+) weak, and (++ and +++) intermediate to strong. RESULTS: AKT protein was associated (p = 0.012) and correlated (p = 0.014; Tau = - 0.288) with the prognostic groups. The immunostaining for TRPM8 (p = 0.010) and NKX3.1 (p = 0.003) proteins differed between malignant tumor and non-tumoral adjacent tissue as well as for proteins in cellular locations (nucleus and cytoplasm). TRPM8 was independently associated with the ISUP grade ≥ 4 (p = 0.024; OR = 8.373; 95% CI = 1.319-53.164). The NKX3.1 showed positive and predominantly strong immunostaining in all patients in both tumoral and non-tumoral adjacent tissues. All metastatic samples had positive immunostaining, with strong intensity for NKX3.1 (p = 0.021; Tau = - 0.302). In the non-metastatic group, this strong protein staining was not observed in any patients. CONCLUSION: This study confirmed that NKX3.1 is highly specific for prostate tissue and indicated that NKX3.1, AKT, and TRPM8 may be candidate markers for prostate cancer prognosis.

Acute infection with Toxoplasma gondii oocysts preferentially activates non-neuronal cells expressing serotonin in the jejunum of rats.

The interaction of Toxoplasma gondii with the gastrointestinal tract of its host is highly regulated. Once ingested, the parasite crosses the epithelium without altering the permeability of the intestinal barrier. Nevertheless, many studies report alterations ranging from structural to functional damage in cells and tissues that make up the wall of the small and large intestine. Although the immune response to the parasite has been extensively studied, the role of serotonin (5-HT) in toxoplasmosis is poorly understood. Here we investigate the distribution of cells expressing 5-HT and its effects on cells and tissues of the jejunal wall of rats after 2, 3, or 7 days of T. gondii infection. KEY RESULTS: Our results show that transposition of the jejunal epithelium by T. gondii leads to ruptures in the basement membrane and activation of the immune system, as confirmed by the decrease in laminin immunostaining and the increase in the number of mast cells, respectively. CONCLUSIONS AND INFERENCES: We showed an increase in the number of enterochromaffin cells and mast cells expressing 5-HT in the jejunal wall. We also observed that the percentage of serotonergic mast cells increased in the total population. Thus, we can suggest that oral infection by T. gondii oocysts preferentially activates non-neuronal cells expressing 5-HT. Together, these results may explain both the changes in the extracellular matrix and the morphology of the enteric ganglia.

Toxoplasma gondii causes increased ICAM-1 and serotonin expression in the jejunum of rats 12 h after infection.

OBJECTIVE: To analyze the remodeling dynamics of total collagen, type I and III, the expression of ICAM-1 and 5-HT in the jejunum of rats. METHODS: Twenty-eight Wistar rats were randomly assigned to two experimental groups: the control group (CG, n = 7) and the infected group (receiving 5,000 sporulated T. gondii oocysts - ME49 strain, genotype II, n = 21). Seven infected rats each at 6 (G6), 12 (G12), and 24 (G24) hours post infection were sacrificed and segments of jejunum were collected for standard histological, histochemical, and immunohistochemistry processing techniques. RESULTS: The infection promoted ICAM-1 and 5-HT expression, type III collagen, and total mast cell increases. However, it also caused a reduction in the area occupied by type I collagen fibers, and in submucosa thickness, and caused ganglion and peri-ganglion alterations. CONCLUSION: The structural damage caused by toxoplasmic infection is intense during the first 24 h post inoculation. At peak dissemination, from 12 to 24 h, there is an increase in ICAM-1 and 5-HT expression, with intense migration of mast cells to the site of infection. There was also a reduction in submucosa thickness, and an effective loss of extracellular matrix (ECM) organization, which included changes in the dynamics of type I and III total collagen deposition.