Phospholipase A2 expression in prostate cancer as a biomarker of good prognosis: A comprehensive study in patients with long follow-up.

BACKGROUND: Phospholipase A2 (PLA2) is a large family of enzymes involved in the inflammatory process that catalyzes the hydrolysis of membrane phospholipids, leading to the production of free fatty acids and lysophospholipids, starting the arachidonic acid cascade. Their expression has been related to the behavior of several cancers. Our objective is to search for PLA2 expression in prostate cancer (PCa) tissue that correlates with prognosis and survival. METHODS: Using qRT-PCR, we analyzed the expression levels of PLA2G1B, PLA2G2A, PLA2G2D, PLA2G4A, PLA2G4B, PLA2G4C, PLA2G4D, PLA2G4E, PLA2G4F, PLA2G6, PLA2G7, PLA2G16, PNPLA1, and PNPLA2 in PCa tissue from 108 patients submitted to radical prostatectomy, followed by a mean time of 163 months. RESULTS: All PLA2 was overexpressed in PCa compared to normal tissue. Interestingly, higher expression of some PLA2 was related to favorable prognostic factors: lower levels of PSA (PLA2G2A, PLA2G4D), lower rates of lymph node metastasis (PLA2G16 and PLA2G1B), and organ-confined disease (PLA2G4A). Most importantly, PLAG4B was independently related to longer disease-free survival. CONCLUSION: This is the first study exploring comprehensively the expression levels of PLA2 in PCa, showing that the higher expression of some PLA2 should be used as biomarkers of good prognosis and longer disease-free survival.

MicroRNA-29b attenuates fibrosis in a rat model of Peyronie's disease.

BACKGROUND: Peyronie's disease is characterized by the formation of fibrotic plaques in the penile tunica albuginea. Effective treatments are limited, warranting the investigation of new promising therapies, such as the application of microRNAs that regulate fibrosis-related genes. OBJECTIVE: We aimed to investigate the therapeutic potential of mimicking microRNA-29b in a fibrin-induced rat model of Peyronie's disease. MATERIAL/METHODS: The study was designed in two phases. To establish an optimal Peyronie's disease model, rats received either human fibrin and thrombin or saline solutions into the tunica albuginea on days 0 and 5. The animal model validation was done through expression and histopathological analyses, the latest by an experienced uropathologist. After validation, we performed microRNA-29b treatment on days 14, 21, and 28 of the study. This phase had control (normal saline) and scramble (microRNA scramble) groups. The mid-penile shaft was removed on day 30 for histological examination and molecular analyses in both study stages. RESULTS: The control group displayed typical tunica albuginea histologic architecture in the animal model validation. In Peyronie's disease group, the Hematoxylin and eosin and Masson Trichrome staining methods demonstrated an interstitial inflammatory process with concomitant dense fibrotic plaques as well as disarrangement of collagen fibers. Additionally, we found out that reduced microRNA-29b (p = 0.05) was associated with significantly increased COL1A1 and transforming growth factor ß1 genes and proteins (p > 0.05) in the Peyronie's disease group. After treatment with mimic microRNA-29b stimulation, the Hematoxylin & eosin and Masson Trichrome staining revealed a discrete and less dense fibrotic plaque. This result was associated with significantly decreasing expression of COL1A1, COL3A1, and transforming growth factor ß1 genes and proteins (p < 0.05). DISCUSSION: The fibrin-induced animal model showed significant histopathological and molecular changes compared to the Control group, suggesting that our model was appropriate. Previous findings have shown that increased expression of microRNA-29b was associated with decreased pathological fibrosis. In the present study, treatment with microRNA-29b decreased the gene and protein expression of collagens and transforming growth factor ß1. This study reveals the therapeutic potential for Peyronie's disease involving molecular targets. CONCLUSION: MicroRNA-29b application on the rat's tunica albuginea attenuated fibrosis, arising as a novel potential strategy for Peyronie's disease management.

Upregulation of shelterin and CST genes and longer telomeres are associated with unfavorable prognostic characteristics in prostate cancer.

INTRODUCTION: Search for new clinical biomarkers targets in prostate cancer (PC) is urgent. Telomeres might be one of these targets. Telomeres are the extremities of linear chromosomes, essential for genome stability and control of cell divisions. Telomere homeostasis relies on the proper functioning of shelterin and CST complexes. Telomeric dysfunction and abnormal expression of its components are reported in most cancers and are associated with PC. Despite this, there are only a few studies about the expression of the main telomere complexes and their relationship with PC progression. We aimed to evaluate the role of shelterin (POT1, TRF2, TPP1, TIN2, and RAP1) and CST (CTC1, STN1, and TEN1) genes and telomere length in the progression of PC. METHODS: We evaluated genetic alterations of shelterin and CST by bioinformatics in samples of localized (n = 499) and metastatic castration-resistant PC (n = 444). We also analyzed the expression of the genes using TCGA (localized PC n = 497 and control n = 152) and experimental approaches, with surgical specimens (localized PC n = 81 and BPH n = 10) and metastatic cell lines (LNCaP, DU145, PC3 and PNT2 as control) by real-time PCR. Real-time PCR also determined the telomere length in the same experimental samples. All acquired data were associated with clinical parameters. RESULTS: Genetic alterations are uncommon in PC, but POT1, TIN2, and TEN1 showed significantly more amplifications in the metastatic cancer. Except for CTC1 and TEN1, which are differentially expressed in localized PC samples, we did not detect an expression pattern relative to control and cell lines. Nevertheless, except for TEN1, the upregulation of all genes is associated with a worse prognosis in localized PC. We also found that increased telomere length is associated with disease aggressiveness in localized PC. CONCLUSION: The upregulation of shelterin and CST genes creates an environment that favors telomere elongation, giving selective advantages for localized PC cells to progress to more aggressive stages of the disease.

Enhancing RECK Expression Through miR-21 Inhibition: A Promising Strategy for Bladder Carcinoma Control.

Bladder carcinoma (BC) is the tenth most frequent malignancy worldwide, with high morbidity and mortality rates. Despite recent treatment advances, high-grade BC and muscle-invasive BC present with significant progression and recurrence rates, urging the need for alternative treatments. The microRNA-21 (miR-21) has superexpression in many malignancies and is associated with cellular invasion and progression. One of its mechanisms of action is the regulation of RECK, a tumor suppressor gene responsible for inhibiting metalloproteinases, including MMP9. In a high-grade urothelial cancer cell line, we aimed to assess if miR-21 downregulation would promote RECK expression and decrease MMP9 expression. We also evaluated cellular migration and proliferation potential by inhibition of this pathway. In a T24 cell line, we inhibited miR-21 expression by transfection of a specific microRNA inhibitor (anti-miR-21). There were also control and scramble groups, the last with a negative microRNA transfected. After the procedure, we performed a genetic expression analysis of miR-21, RECK, and MMP9 through qPCR. Migration, proliferation, and protein expression were evaluated via wound healing assay, colony formation assay, flow cytometry, and immunofluorescence.After anti-miR-21 transfection, miR-21 expression decreased with RECK upregulation and MMP9 downregulation. The immunofluorescence assay showed a significant increase in RECK protein expression (p < 0.0001) and a decrease in MMP9 protein expression (p = 0.0101). The anti-miR-21 transfection significantly reduced cellular migration in the wound healing assay (p < 0.0001). Furthermore, in the colony formation assay, the anti-miR-21 group demonstrated reduced cellular proliferation (p = 0.0008), also revealed in the cell cycle analysis by flow cytometry (p = 0.0038). Our results corroborate the hypothesis that miR-21 is associated with BC cellular migration and proliferation, revealing its potential as a new effective treatment for this pathology.

The Effect of Gene Editing by CRISPR-Cas9 of miR-21 and the Indirect Target MMP9 in Metastatic Prostate Cancer.

Prostate cancer (PCa) has a high prevalence and represents an important health problem, with an increased risk of metastasis. With the advance of CRISPR-Cas9 genome editing, new possibilities have been created for investigating PCa. The technique is effective in knockout oncogenes, reducing tumor resistance. MMP9 and miR-21 target genes are associated with PCa progression; therefore, we evaluated the MMP-9 and miR-21 targets in PCa using the CRISPR-Cas9 system. Single guide RNAs (sgRNAs) of MMP9 and miR-21 sequences were inserted into a PX-330 plasmid, and transfected in DU145 and PC-3 PCa cell lines. MMP9 and RECK expression was assessed by qPCR, WB, and IF. The miR-21 targets, integrins, BAX and mTOR, were evaluated by qPCR. Flow cytometry was performed with Annexin5, 7-AAD and Ki67 markers. Invasion assays were performed with Matrigel. The miR-21 CRISPR-Cas9-edited cells upregulated RECK, MARCKS, BTG2, and PDCD4. CDH1, ITGB3 and ITGB1 were increased in MMP9 and miR-21 CRISPR-Cas9-edited cells. Increased BAX and decreased mTOR were observed in MMP9 and miR-21 CRISPR-Cas9-edited cells. Reduced cell proliferation, increased apoptosis and low invasion in MMP9 and miR-21 edited cells was observed, compared to Scramble. CRISPR-Cas9-edited cells of miR-21 and MMP9 attenuate cell proliferation, invasion and stimulate apoptosis, impeding PCa evolution.

Overexpression of miR-17-5p may negatively impact p300/CBP factor-associated inflammation in a hypercholesterolemic advanced prostate cancer model.

BACKGROUND: Previously, we demonstrated that cholesterol triggers the increase in p300/CBP-associated factor (PCAF), targeted by miR-17-5p. The p300, IL-6, PCAF, and miR-17-5p genes have important and contradictory roles in inflammation and prostate cancer (PCa). This study aimed to demonstrate the potential anti-inflammatory effect of miR-17-5 in an advanced PCa model with diet-induced hypercholesterolemia. METHODS AND RESULTS: In vitro, using the PC-3 cell line, we show that induction of miR-17-5p reduces p300 and PCAF expression, increases apoptosis, and decreases cell migration. Furthermore, we demonstrate that supplementing this same cell with cholesterol (2 µg/mL) triggers increased p300, IL-6, and PCAF. In vivo, after establishing the hypercholesterolemic (HCOL) model, xenografts were treated with miR-17-5p. Increased expression of this miR after intratumoral injections attenuated tumor growth in the control and HCOL animals and reduced cell proliferation. CONCLUSION: Our results demonstrate that inducing miR-17-5p expression suppresses tumor growth and inflammatory mediator expression. Further studies should be conducted to fully explore the role of miR-17-5p and the involvement of inflammatory mediators p300, PCAF, and IL-6.

Intratumoral Restoration of miR-137 Plus Cholesterol Favors Homeostasis of the miR-137/Coactivator p160/AR Axis and Negatively Modulates Tumor Progression in Advanced Prostate Cancer.

MicroRNAs (miRNAs) have gained a prominent role as biomarkers in prostate cancer (PCa). Our study aimed to evaluate the potential suppressive effect of miR-137 in a model of advanced PCa with and without diet-induced hypercholesterolemia. In vitro, PC-3 cells were treated with 50 pmol of mimic miR-137 for 24 h, and gene and protein expression levels of SRC-1, SRC-2, SRC-3, and AR were evaluated by qPCR and immunofluorescence. We also assessed migration rate, invasion, colony-forming ability, and flow cytometry assays (apoptosis and cell cycle) after 24 h of miRNA treatment. For in vivo experiments, 16 male NOD/SCID mice were used to evaluate the effect of restoring miR-137 expression together with cholesterol. The animals were fed a standard (SD) or hypercholesterolemic (HCOL) diet for 21 days. After this, we xenografted PC-3 LUC-MC6 cells into their subcutaneous tissue. Tumor volume and bioluminescence intensity were measured weekly. After the tumors reached 50 mm3, we started intratumor treatments with a miR-137 mimic, at a dose of 6 µg weekly for four weeks. Ultimately, the animals were killed, and the xenografts were resected and analyzed for gene and protein expression. The animals' serum was collected to evaluate the lipid profile. The in vitro results showed that miR-137 could inhibit the transcription and translation of the p160 family, SRC-1, SRC-2, and SRC-3, and indirectly reduce the expression of AR. After these analyses, it was determined that increased miR-137 inhibits cell migration and invasion and impacts reduced proliferation and increased apoptosis rates. The in vivo results demonstrated that tumor growth was arrested after the intratumoral restoration of miR-137, and proliferation levels were reduced in the SD and HCOL groups. Interestingly, the tumor growth retention response was more significant in the HCOL group. We conclude that miR-137 is a potential therapeutic miRNA that, in association with androgen precursors, can restore and reinstate the AR-mediated axis of transcription and transactivation of androgenic pathway homeostasis. Further studies involving the miR-137/coregulator/AR/cholesterol axis should be conducted to evaluate this miR in a clinical context.

Cholesterol Triggers Nuclear Co-Association of Androgen Receptor, p160 Steroid Coactivators, and p300/CBP-Associated Factor Leading to Androgenic Axis Transactivation in Castration-Resistant Prostate Cancer.

BACKGROUND/AIMS: Cholesterol modulates intratumoral androgenic signaling in prostate cancer; however, the molecular mechanisms underlying these changes in castration-resistant prostate cancer (CRPC) are not fully elucidated. Herein, we investigated the effect of cholesterol on androgen receptor (AR) coactivators expression and tumorigenesis in vitro and in vivo. METHODS: Herein, we monitored the expression of AR coactivators (SRC-1, 2, 3 and PCAF) genes in PC-3 cells exposed to 2µg/mL of cholesterol for 8 hours by qPCR. We also performed cell migration at 0, 8, 24, 48 and 72h and flow cytometry assays (viability, apoptosis, and cell cycle) after a 24h exposure. Immunofluorescence assay was performed to evaluate the protein expression of the AR coactivators. Additionally, in vivo experiments were conducted using 22 male NOD/SCID mice. Mice were fed a standard (Control) or hypercholesterolemic (HCOL) diet for 21 days and then subcutaneously implanted with PC-3 cells. The tumor volume was calculated every two days, and after four weeks, the tumors were resected, weighed, and the serum lipid profile was measured. We also measured the intratumoral lipid profile and AR coactivators gene and protein expression by qPCR and Western Blot, respectively. Intratumor testosterone and dihydrotestosterone (DHT) concentrations were determined using ELISA. RESULTS: Cholesterol up-regulated the gene expression of coactivators SRC-1, SRC-2, SRC-3and PCAF, increasing AR expression in PC-3 cells. Next, cholesterol-supplemented PC-3 cells exhibited increased cell migration and altered cell cycle phases, leading to changes in proliferation and reduced apoptosis. We found that SRC-1, SRC-2, SRC-3 and PCAF proteins co-localized in the nucleus of cholesterol-supplemented cells and co-associate with AR. In the in vivo model, the hypercholesterolemic (HCOL) group displayed higher serum total and intratumoral cholesterol levels, increased testosterone and dihydrotestosterone concentrations, and up-regulated AR coactivator expression. The tumor volume of the HCOL group was significantly higher than the control group. CONCLUSION: Our findings revealed that increased nuclear translocation of the coactivators leads to up-regulated AR gene and protein expression, potentially influencing tumor progression. Studies targeting cholesterol-modulated changes in AR coactivator expression may provide insights into the molecular mechanisms associated with the CRPC phenotype.

Can we use Ki67 expression to predict prostate cancer aggressiveness?

INTRODUCTION: specialists have an urge for biomarkers that can discriminate indolent prostate cancer from aggressive tumors. Ki67 is a proliferation marker, and its expression is associated with the aggressiveness of several cancers. OBJECTIVE: analyze the expression of Ki67 in prostate cancer samples correlating with the aggressiveness of the disease. METHODS: Ki67 mRNA levels were determined utilizing data from a TCGA cohort (Tumor(n)=492 and control(n)=52). The protein expression was determined on 94 biopsies from patients by immunohistochemical assay. RESULTS: in mRNA, the Ki67 upregulation is associated with cancer tissue (p<0.0001) and worst disease-free survival (p=0.035). The protein upregulation is associated with increase of the ISUP score (p<0.0001), cancer stage (p=0.05), biochemical recurrence (p=0.0006) and metastasis (p<0.0001). We also show a positive correlation between Ki67 expression and ISUP score (r=0.5112, p<0.0001) and disease risk stratification (r=0.3388, p=0.0009). Ki67 expression is a factor independently associated with biochemical recurrence (p=0.002) and metastasis (p<0.0001). Finally, the patients with high Ki67expression shows better survival regarding biochemical recurrence (p=0.008) and metastasis (p=0.056). Patients with high Ki67 expression are 2.62 times more likely to develop biochemical recurrence (p=0.036). CONCLUSION: Ki67 upregulation is associated with prostate cancer aggressiveness.

Combined spinal and general anesthesia attenuate tumor promoting effects of surgery. An experimental animal study.

Background: Radical prostatectomy, a standard management approach for localized Prostate Cancer (PC), may cause a stress response associated with immune modulating effects. Regional anesthesia was hypothesized to reduce the immune effects of surgery by minimizing the neuroendocrine surgical stress response, thus mitigating tumor cells dissemination. Our primary objective was to investigate whether the use of spinal blocks attenuates PC tumor cells dissemination on an animal model. We also assessed the number of circulating NK cells and the amount of inflammatory and anti-inflammatory cytokines. Materials and methods: A subcutaneous tumor model, with PC-3M cell line transfected with a luciferase-producing gene (PC-3M-luc-C6) was used. After proper tumor establishment and before tumors became metastatic, animals were submitted to tumor excision surgeries under general or combined (general and spinal) anesthesia. A control group was only anesthetized with general anesthesia. Results: The subcutaneous tumor model with PC-3M-luc-C6 cells was effective in causing distant metastasis after 35 days. The number of circulating tumor cells increased in animals that underwent surgery under general anesthesia alone compared to the group submitted to combined anesthesia. Interleukin 6 levels were different in all groups, with increase in the general anesthesia group. Conclusion: Our results suggest that combination of spinal and general anesthesia may attenuate the suppression of innate tumor immunity and it might be related to a reduction in the neuroendocrine response to surgery. Institutional protocol number: Animal Ethics Committee 1332/2019.

Pan-cancer analysis reveals that CTC1-STN1-TEN1 (CST) complex may have a key position in oncology.

Telomere dysfunction is one of the hallmarks of cancer, which puts telomere-associated genes in a prominent position in oncology. The CTC1-STN1-TEN1 (CST) complex is vital for telomere maintenance and participates in several steps of DNA metabolism, such as repair and replication, essential functions for malignant cells. Despite this, little is known about these genes in cancer biology. Here, using bioinformatics tools, we performed a study in 33 cancer types and over 10,000 TCGA samples analyzing the role of the CST complex in cancer. We obtained the somatic landscape and gene expression patterns of each of the subunits of the complex studied. Furthermore, we show that CST is important for genetic stability and nucleic acid metabolism in cancer. We identify possible interactors, transcription factors, and microRNAs associated with CST and two drugs that may disrupt their pathways. In addition, we show that CST gene expression is associated with cancer survival and recurrence in several tumor types. Finally, we show negative and positive correlations between immune checkpoint genes and CST in different types of cancer. With this work, we corroborate the importance of these genes in cancer biology and open perspectives for their use in other works in the field.

First detection of Feline morbillivirus infection in white-eared opossums (Didelphis albiventris, Lund, 1840), a non-feline host.

Feline Morbillivirus (FeMV) was first detected in 2012 in domestic cats from Hong Kong and was found to be associated with tubulointerstitial nephritis and chronic kidney disease. In subsequent studies in other countries, FeMV was detected in asymptomatic cats. However, it is not clear whether FeMV plays a role as a pathogen in the kidney diseases of cats, and other epidemiological data are still unknown. To date, studies have reported the presence of FeMV exclusively in domestic cats. This study is the first molecular detection of the FeMV RNA associated with pathological and immunohistochemical findings in a synanthropic marsupial, the white-eared opossum (Didelphis albiventris), inhabiting peri-urban areas of north-central Parana, Southern Brazil. Molecular techniques identified the viral RNA in the lungs and kidneys. Histopathologic evaluation of these tissues revealed interstitial pneumonia in the lungs with lymphocytic nephritis and tubular necrosis in the kidneys. Immunohistochemistry assays detected positive intralesional immunoreactivity to N protein of FeMV within the lungs and kidneys. A FeMV opossum strain was isolated in Crandell Rees feline kidney lineage cells, resulting in syncytia formation and cell death. Therefore, these results support the ability of FeMV to infect other mammal species and reinforce the possibility of the opossum to be a disseminator of this virus among domestic and wild animals.

Downregulation of miR-29b is associated with Peyronie's disease.

BACKGROUND: Peyronie's disease (PD) is characterized by the formation of fibrous plaque in tunica albuginea, causing several problems in patients. The etiology of this disease is not fully understood, and there are few effective treatments. To better understand the molecular pathways of PD, we studied miR-29b, a microRNA that could be involved with this illness. MicroRNAs are endogenous molecules that act by inhibiting messenger RNA. MiR-29b regulates 11 of 20 collagen genes and the TGF-ß1 gene, which are related to PD progression. METHODS: We compared miR-29b expression in 11 patients with PD and 14 patients without PD (control group). For the patients with PD, we utilized samples from the fibrous plaque (n = 9), from the tunica albuginea (n = 11), and from the corpus cavernosum (n = 8). For the control group, we utilized samples from the tunica albuginea (n = 14) and from the corpus cavernosum (n = 10). MiR-29b expression was determined by q-PCR. RESULTS: We found a downregulation of miR-29b in the fibrous plaque, tunica albuginea and corpus cavernosum of patients with PD in comparison with the control group (p = 0.0484, p = 0.0025, and p = 0.0016, respectively). CONCLUSION: Although our study has a small sample, we showed for the first time an evidence that the downregulation of miR-29b is associated with PD.

Can we use Ki67 expression to predict prostate cancer aggressiveness? / Podemos usar a expressão de Ki67 para prever a agressividade do câncer de próstata?

ABSTRACT Introduction: specialists have an urge for biomarkers that can discriminate indolent prostate cancer from aggressive tumors. Ki67 is a proliferation marker, and its expression is associated with the aggressiveness of several cancers. Objective: analyze the expression of Ki67 in prostate cancer samples correlating with the aggressiveness of the disease. Methods: Ki67 mRNA levels were determined utilizing data from a TCGA cohort (Tumor(n)=492 and control(n)=52). The protein expression was determined on 94 biopsies from patients by immunohistochemical assay. Results: in mRNA, the Ki67 upregulation is associated with cancer tissue (p<0.0001) and worst disease-free survival (p=0.035). The protein upregulation is associated with increase of the ISUP score (p<0.0001), cancer stage (p=0.05), biochemical recurrence (p=0.0006) and metastasis (p<0.0001). We also show a positive correlation between Ki67 expression and ISUP score (r=0.5112, p<0.0001) and disease risk stratification (r=0.3388, p=0.0009). Ki67 expression is a factor independently associated with biochemical recurrence (p=0.002) and metastasis (p<0.0001). Finally, the patients with high Ki67expression shows better survival regarding biochemical recurrence (p=0.008) and metastasis (p=0.056). Patients with high Ki67 expression are 2.62 times more likely to develop biochemical recurrence (p=0.036). Conclusion: Ki67 upregulation is associated with prostate cancer aggressiveness.

RESUMO Introdução: especialistas precisam biomarcadores que podem discriminar o câncer de próstata indolente de tumores agressivos. Ki67 é um marcador de proliferação, e sua expressão está associada à agressividade de vários tumores. Objetivo: analisar a expressão do Ki67 em amostras de câncer de próstata correlacionando com a agressividade da doença. Métodos: os níveis de mRNA de Ki67 foram determinados utilizando dados de uma coorte de TCGA (Tumor(n)=492 e controle(n)=52). A expressão da proteína foi determinada em 94 biópsias de pacientes por ensaio imuno-histoquímica. Resultados: no mRNA, a superexpressão Ki67 está associada ao tecido canceroso (p<0,0001) e à pior sobrevida livre de doença (p=0,035). A superexpressão proteica está associada ao aumento do escore ISUP (p<0,0001), estágio de câncer (p=0,05), recorrência bioquímica (p=0,0006) e metástase (p<0,0001). Também mostramos uma correlação positiva entre a expressão Ki67 e o escore ISUP (r=0,5112, p<0,0001) e a estratificação de risco de doença (r=0,3388, p=0,0009). A expressão Ki67 é um fator independentemente associado à recorrência bioquímica (p=0,002) e metástase (p<0,0001). Finalmente, os pacientes com alta expressão de Ki67 expression mostram melhor sobrevivência em relação à recorrência bioquímica (p=0,008) e metástase (p=0,056). Os pacientes com alta expressão de Ki67 são 2,62 vezes mais propensos a desenvolver recorrência bioquímica (p=0,036). Conclusão: a superexpressão Ki67 está associada à agressividade do câncer de próstata.

Can increased expression of miR-Let-7c reduce the transition potential of high-grade urothelial carcinoma?

BACKGROUND: Bladder cancer is the leading transitional cell carcinoma affecting men and women with high morbidity and mortality rates, justifying the need to develop new molecular target therapies using microRNAs. This study aimed to evaluate the behavior of the T24 cell line after transfection with miR-Let-7c precursor mimic through invasion, migration, apoptosis, and cell cycle assays. METHODS AND RESULTS: T24 cell was transfected with the Let-7c mimic and its respective control and evaluated after 24 h. The expression levels of miR-Let-7c were analyzed by qPCR. We performed wound healing, Matrigel and flow cytometry, apoptosis, and cell cycle assays to determine its effect on cellular processes. Cells transfected with miR-Let-7c showed increased apoptosis rates (p = 0.019), decreased migration 24 h (p = 0.031) and 48 h (p = 0.0006), invasion potential (p = 0.0007), and cell proliferation (p = 0.002). CONCLUSIONS: Our results demonstrate that miR-Let-7c can act in different pathways of the carcinogenic cellular processes of muscle-invasive urothelial carcinoma cells, inhibiting cell proliferation and increasing apoptosis levels, consequently limiting their invasion potential. However, further studies should be carried out better to elucidate this microRNA's role in high-grade urothelial carcinomas and unveil which targets this microRNA may present, which are intrinsically related to the cancer survival pathways.

Telomeric zinc-finger associated protein (TZAP) in cancer biology: friend or foe?

The new identified protein telomeric zinc-finger associated protein (TZAP) is a negative regulator of telomere length. Since telomere length and telomere maintenance mechanisms are essential to cancer progression, TZAP is considered a new player in cancer biology. Here we aimed to analyze TZAP using the Cancer Genome Atlas data in a Pan-Cancer approach. We gathering data from TCGA Pan-Cancer studies utilizing cBioPortal, GEPIA and UALCAN. In total we analyzed 33 types of cancer (n=9664) and their respective controls (n=711). TZAP is transcribed in all cancers but less than 5% of all tumors show any somatic changes. TZAP was downregulated in kidney chromophobe carcinoma, and upregulated in esophageal cancer, head and neck squamous cell carcinomas, kidney renal clear cell carcinoma and in liver hepatocellular carcinoma. Globally, TZAP expression is related to favorable prognosis, associated to better overall and disease-free survival. Looking to specific tumors, TZAP expression has a dual behavior. Its downregulation is associated with poor prognosis in cervical squamous cell carcinoma, in kidney renal clear cell carcinoma, kidney papillary cell carcinoma, lung adenocarcinoma and pancreas adenocarcinoma. On the contrary, in adrenocortical carcinoma, colon and rectal cancer, brain lower grade glioma and prostate adenocarcinoma the upregulation of TZAP is related with poor prognosis. TZAP expression has a positive correlation with TRF1 and TRF2 in normal tissue but not in cancer. Our analyses indicate that TZAP has an important role in oncology and may be considered as a potential biomarker.

MiR-200c-3p expression may be associated with worsening of the clinical course of patients with COVID-19.

COVID-19 represents a public health emergency, whose mechanism of which is not fully understood. It is speculated that microRNAs may play a crucial role in host cells after infection by SARS-CoV-2. Thus, our study aimed to analyze the expression of miR-200c-3p in saliva samples from patients with COVID-19. One handred eleven samples from patients with COVID-19 were divided into 4 groups. Group I: 39 patients negative for Covid-19; Group II: 37 positive and symptomatic patients, with no indication of hospitalization; Group III: 21 patients with respiratory disorders (hospitalized); Group IV: 14 patients with severe conditions (oxygen therapy). The expression levels of miR-200c-3p were determined using qPCR. We found greater expression of miR-200c-3p in patients in group IV (p<0.0001), and also verified that patients aged ≥42 years had a higher expression of this miR (p=0.013). Logistic regression analysis revealed that the expression of miR-200c-3p and systemic arterial hypertension are factors independently associated with patients in group IV (p<0.0001). Our results suggest that miR-200c-3p is a predictor of severity independent of COVID-19 risk factors, which could represent a way of screening patients affected by SARS-CoV-2.

Prognostic value of TERF1 expression in prostate cancer.

BACKGROUND: Telomere dysfunction is one of the hallmarks of cancer and is crucial to prostate carcinogenesis. TERF1 is a gene essential to telomere maintenance, and its dysfunction has already been associates with several cancers. TERF1 is a target of miR-155, and this microRNA can inhibit its expression and promotes carcinogenesis in breast cancer. We aim to analyze TERF1, in gene and mRNA level, involvement in prostate cancer progression. RESULTS: Alterations in TERF1 DNA were evaluated using datasets of primary tumor and castration-resistant tumors (CRPC) deposited in cBioportal. The expression of TERF1 mRNA levels was assessed utilizing TCGA datasets, clinical specimens, and metastatic prostate cancer cell lines (LNCaP, DU145, and PC3). Six percent of localized prostate cancer presents alterations in TERF1 (the majority of that was amplifications). In the CRPC cohort, 26% of samples had TERF1 amplification. Patients with TERF1 alterations had the worst overall survival only on localized cancer cohort (p = 0.0027). In the TCGA cohort, mRNA levels of TERF1 were downregulated in comparison with normal tissue (p = 0.0013) and upregulated in tumors that invade lymph nodes (p = 0.0059). The upregulation of TERF1 is also associated with worst overall survival (p = 0.0028) and disease-free survival (p = 0.0023). There is a positive correlation between TERF1 and androgen receptor expression in cancer tissue (r = 0.53, p < 0.00001) but not on normal tissue (r = - 0.16, p = 0.12). In the clinical specimens, there is no detectable expression of TERF1 and upregulation of miR-155 (p = 0.0348). In cell lines, TERF1 expression was higher in LNCaP and was progressively lower in DU145 and PC3 (p = 0.0327) with no differences in miR-155 expression. CONCLUSION: Amplification/upregulation of TERF1 was associated with the worst prognostic in localized prostate cancer. Our results corroborate that miR-155 regulates TERF1 expression in prostate cancer. TERF1 has the potential to become a biomarker in prostate cancer.

Shorter leukocyte telomere length is associated with severity of COVID-19 infection.

The infection by COVID-19 is a serious global public health problem. An efficient way to improve this disease's clinical management would be to characterize patients at higher risk of progressing to critically severe infection using prognostic biomarkers. The telomere length could be used for this purpose. Telomeres are responsible for controlling the number of maximum cell divisions. The telomere length is a biomarker of aging and several diseases. We aimed to compare leukocyte telomere length (LTL) between patients without COVID-19 and patients with different clinical severity of the infection. Were included 53 patients who underwent SARS-CoV-2 PCR divided in four groups. The first group was composed by patients with a negative diagnosis for COVID-19 (n = 12). The other three groups consisted of patients with a confirmed diagnosis of COVID-19 divided according to the severity of the disease: mild (n = 15), moderate (n = 17) and severe (n = 9). The LTL was determined by Q-PCR. The severe group had the shortest LTL, followed by the moderate group. The negative and mild groups showed no differences. There is an increase of patients with hypertension (p = 0.0099) and diabetes (p = 0.0067) in moderate and severe groups. Severe group was composed by older patients in comparison with the other three groups (p = 0.0083). Regarding sex, there was no significant difference between groups (p = 0.6279). In an ordinal regression model, only LTL and diabetes were significantly associated with disease severity. Shorter telomere length was significantly associated with the severity of COVID-19 infection, which can be useful as a biomarker or to better understand the SARS-CoV-2 pathophysiology.

A novel supplement with yeast ß-glucan, prebiotic, minerals and Silybum marianum synergistically modulates metabolic and inflammatory pathways and improves steatosis in obese mice.

OBJECTIVE: To evaluate the synergic effects of a novel oral supplement formulation, containing prebiotics, yeast ß-glucans, minerals and silymarin (Silybum marianum), on lipid and glycidic metabolism, inflammatory and mitochondrial proteins of the liver, in control and high-fat diet-induced obese mice. METHODS: After an acclimation period, 32 male C57BL/6 mice were divided into the following groups: nonfat diet (NFD) vehicle, NFD supplemented, high-fat diet (HFD) vehicle and HFD supplemented. The vehicle and experimental formulation were administered orally by gavage once a day during the last four weeks of the diet (28 consecutive days). We then evaluated energy homeostasis, inflammation, and mitochondrial protein expression in these groups of mice. RESULTS: After four weeks of supplementation, study groups experienced reduced glycemia, dyslipidemia, fat, and hepatic fibrosis levels. Additionally, proliferator-activated receptor-α, AMP-activated protein kinase-1α, peroxisome proliferator-activated receptor γ co-activator-1α, and mitochondrial transcription factor A expression levels were augmented; however, levels of inhibitor of nuclear factor-κB kinase subunit α and p65 nuclear factor-κB expression, and oxidative markers were reduced. Notably, the cortisol/C-reactive protein ratio, a well-characterized marker of the hypothalamic-pituitary-adrenal axis immune interface status, was found to be modulated by the supplement. CONCLUSION: We discovered that the novel supplement was able to modify different antioxidant, metabolic and inflammatory pathways, improving the energy homeostasis and inflammatory status, and consequently alleviated hepatic steatosis.