Role of a Mixture of Polyphenol Compounds Released after Blueberry Fermentation in Chemoprevention of Mammary Carcinoma: In Vivo Involvement of miR-145.

Epigenetic mechanisms such as microRNA (miRNA) deregulation seem to exert a central role in breast cancer initiation and progression. Therefore, targeting epigenetics deregulation may be an effective strategy for preventing and halting carcinogenesis. Studies have revealed the significant role of naturally occurring polyphenolic compounds derived from fermented blueberry fruits in cancer chemoprevention by modulation of cancer stem cell development through the epigenetic mechanism and regulation of cellular signaling pathways. In this study, we first investigated the phytochemical changes during the blueberry fermentation process. Fermentation favored the release of oligomers and bioactive compounds such as protocatechuic acid (PCA), gallic acid, and catechol. Next, we investigated the chemopreventive potentials of a polyphenolic mixture containing PCA, gallic acid, and catechin found in fermented blueberry juice in a breast cancer model by measuring miRNA expression and the signaling pathways involved in breast cancer stemness and invasion. To this end, 4T1 and MDA-MB-231 cell lines were treated with different doses of the polyphenolic mixture for 24 h. Additionally, female Balb/c mice were fed with this mixture for five weeks; two weeks before and three weeks after receiving 4T1 cells. Mammosphere formation was assayed in both cell lines and the single-cell suspension obtained from the tumor. Lung metastases were counted by isolating 6-thioguanine-resistant cells present in the lungs. In addition, we conducted RT-qPCR and Western blot analysis to validate the expression of targeted miRNAs and proteins, respectively. We found a significant reduction in mammosphere formation in both cell lines treated with the mixture and in tumoral primary cells isolated from mice treated with the polyphenolic compound. The number of colony-forming units of 4T1 cells in the lungs was significantly lower in the treatment group compared to the control group. miR-145 expression significantly increased in the tumor samples of mice treated with the polyphenolic mixture compared to the control group. Furthermore, a significant increase in FOXO1 levels was noted in both cell lines treated with the mixture. Overall, our results show that phenolic compounds found in fermented blueberry delay the formation of tumor-initiating cells in vitro and in vivo and reduce the spread of metastatic cells. The protective mechanisms seem to be related, at least partly, to the epigenetic modulation of mir-145 and its signaling pathways.

Lentinula edodes Cultured Extract and Rouxiella badensis subsp. acadiensis (Canan SV-53) Intake Alleviates Immune Deregulation and Inflammation by Modulating Signaling Pathways and Epigenetic Mechanisms.

Puberty is a critical developmental period of life characterized by marked physiological changes, including changes in the immune system and gut microbiota development. Exposure to inflammation induced by immune stressors during puberty has been found to stimulate central inflammation and lead to immune disturbance at distant sites from the gut; however, its enduring effects on gut immunity are not well explored. Therefore, in this study, we used a pubertal lipopolysaccharides (LPS)-induced inflammation mouse model to mimic pubertal exposure to inflammation and dysbiosis. We hypothesized that pubertal LPS-induced inflammation may cause long-term dysfunction in gut immunity by enduring dysregulation of inflammatory signaling and epigenetic changes, while prebiotic/probiotic intake may mitigate the gut immune system deregulation later in life. To this end, four-week-old female Balb/c mice were fed prebiotics/probiotics and exposed to LPS in the pubertal window. To better decipher the acute and enduring immunoprotective effects of biotic intake, we addressed the effect of treatment on interleukin (IL)-17 signaling related-cytokines and pathways. In addition, the effect of treatment on gut microbiota and epigenetic alterations, including changes in microRNA (miRNA) expression and DNA methylation, were studied. Our results revealed a significant dysregulation in selected cytokines, proteins, and miRNAs involved in key signaling pathways related to IL-17 production and function, including IL-17A and F, IL-6, IL-1ß, transforming growth factor-ß (TGF-ß), signal transducer and activator of transcription-3 (STAT3), p-STAT3, forkhead box O1 (FOXO1), and miR-145 in the small intestine of adult mice challenged with LPS during puberty. In contrast, dietary interventions mitigated the lasting adverse effects of LPS on gut immune function, partly through epigenetic mechanisms. A DNA methylation analysis demonstrated that enduring changes in gut immunity in adult mice might be linked to differentially methylated genes, including Lpb, Rorc, Runx1, Il17ra, Rac1, Ccl5, and Il10, involved in Th17 cell differentiation and IL-17 production and signaling. In addition, prebiotic administration prevented LPS-induced changes in the gut microbiota in pubertal mice. Together, these results indicate that following a healthy diet rich in prebiotics and probiotics is an optimal strategy for programming immune system function in the critical developmental windows of life and controlling inflammation later in life.

Prevalence of metabolically unhealthy obesity, overweight, and normal weight and the associated risk factors in a southern coastal region, Iran (the PERSIAN cohort study): a cross-sectional study.

BACKGROUND: Obesity is a substantial risk factor for cardiovascular and metabolic diseases. Epidemiologic studies have shown that some obese and overweight individuals are metabolically healthy. We aimed to determine the prevalence of metabolically unhealthy obesity (MUO), metabolically unhealthy overweight (MUOW), and metabolically unhealthy normal weight (MUNW) in a southern coastal area of Iran, Bandare-Kong Non-Communicable Diseases (BKNCD) Cohort Study. METHODS: This population-based study included the participants of BKNCD, as part of the Prospective Epidemiological Research Studies in IrAN (PERSIAN). Metabolic health was defined as not fulfilling the metabolic syndrome (MetS) criteria. RESULTS: Among the 3917 participants in this study with the mean age of 48.29 ± 9.39 years, including 1691 (43.2%) males, the age- and sex-standardized prevalence of MUO, MUOW, and MUNW was 13.9, 16.8, and 6.4%, respectively. Binary logistic regression analysis revealed that the adjusted odds of all metabolically unhealthy states were higher in older age groups, except for MUO whose adjusted odds were lower in the 65-70 age group compared to the 55-64 age group. Illiteracy was significantly correlated with MUOW (adjusted OR: 1.43, 95% CI 1.09-1.87, P = 0.010); however, it was not associated with MUO or MUNW. Higher body mass index (BMI) was significantly correlated with MUNW but it was not associated with MUOW or MUO. Higher waist circumference (WC) was also significantly associated with all metabolically unhealthy states. CONCLUSION: The age- and sex-standardized prevalence of MUO, MUOW, and MUNW was 13.9, 16.8, and 6.4%, respectively in the current study. Advanced age and higher WC were significantly correlated with all metabolically unhealthy states, while illiteracy and higher BMI were only associated with MUOW and MUNW, respectively. Metabolic health rather than weight loss should be the focus and objective of public prevention programs.

Polyphenol-Enriched Blueberry Preparation Controls Breast Cancer Stem Cells by Targeting FOXO1 and miR-145.

Scientific evidence supports the early deregulation of epigenetic profiles during breast carcinogenesis. Research shows that cellular transformation, carcinogenesis, and stemness maintenance are regulated by epigenetic-specific changes that involve microRNAs (miRNAs). Dietary bioactive compounds such as blueberry polyphenols may modulate susceptibility to breast cancer by the modulation of CSC survival and self-renewal pathways through the epigenetic mechanism, including the regulation of miRNA expression. Therefore, the current study aimed to assay the effect of polyphenol enriched blueberry preparation (PEBP) or non-fermented blueberry juice (NBJ) on the modulation of miRNA signature and the target proteins associated with different clinical-pathological characteristics of breast cancer such as stemness, invasion, and chemoresistance using breast cancer cell lines. To this end, 4T1 and MB-MDM-231 cell lines were exposed to NBJ or PEBP for 24 h. miRNA profiling was performed in breast cancer cell cultures, and RT-qPCR was undertaken to assay the expression of target miRNA. The expression of target proteins was examined by Western blotting. Profiling of miRNA revealed that several miRNAs associated with different clinical-pathological characteristics were differentially expressed in cells treated with PEBP. The validation study showed significant downregulation of oncogenic miR-210 expression in both 4T1 and MDA-MB-231 cells exposed to PEBP. In addition, expression of tumor suppressor miR-145 was significantly increased in both cell lines treated with PEBP. Western blot analysis showed a significant increase in the relative expression of FOXO1 in 4T1 and MDA-MB-231 cells exposed to PEBP and in MDA-MB-231 cells exposed to NBJ. Furthermore, a significant decrease was observed in the relative expression of N-RAS in 4T1 and MDA-MB-231 cells exposed to PEBP and in MDA-MB-231 cells exposed to NBJ. Our data indicate a potential chemoprevention role of PEBP through the modulation of miRNA expression, particularly miR-210 and miR-145, and protection against breast cancer development and progression. Thus, PEBP may represent a source for novel chemopreventative agents against breast cancer.

Probiotics in Treatment of Viral Respiratory Infections and Neuroinflammatory Disorders.

Inflammation is a biological response to the activation of the immune system by various infectious or non-infectious agents, which may lead to tissue damage and various diseases. Gut commensal bacteria maintain a symbiotic relationship with the host and display a critical function in the homeostasis of the host immune system. Disturbance to the gut microbiota leads to immune dysfunction both locally and at distant sites, which causes inflammatory conditions not only in the intestine but also in the other organs such as lungs and brain, and may induce a disease state. Probiotics are well known to reinforce immunity and counteract inflammation by restoring symbiosis within the gut microbiota. As a result, probiotics protect against various diseases, including respiratory infections and neuroinflammatory disorders. A growing body of research supports the beneficial role of probiotics in lung and mental health through modulating the gut-lung and gut-brain axes. In the current paper, we discuss the potential role of probiotics in the treatment of viral respiratory infections, including the COVID-19 disease, as major public health crisis in 2020, and influenza virus infection, as well as treatment of neurological disorders like multiple sclerosis and other mental illnesses.

Development of a Modified Friedewald's Formula to Calculate Low-Density Lipoprotein in an Iranian Population.

Background: Elevated low-density lipoprotein cholesterol (LDL-C) is a significant risk factor for cardiovascular diseases. LDL-C can be directly measured using various methods, but this requires expensive equipment. Currently, clinical laboratories estimate LDL-C based on Friedewald's formula (FF). We aimed to develop a modified formula based on directly measured LDL-C (D-LDL-C) values in a large population in Southern Iran and compare the results with various other estimation formulas. Methods: The participants of this cross-sectional study were adults aged >18 years living in Southern Iran. Blood samples from 15,200 individuals were collected, and the measured lipid parameters were randomly divided into training (n=10,184) and validation (n=5,016) datasets. A new formula was developed using a linear regression model, and its accuracy was validated. Pearson's correlation and Cohen's kappa were used to determin the relationship between D-LDL-C and calculated LDL-C (C-LDL-C). Results: The developed formula for the estimation of LDL-C was 0.857 total cholesterol (TC)-0.915 high-density lipoprotein cholesterol (HDL-C)-0.115 triglycerides (TG). Based on our proposed formula, for TG<150 and TG≥150 mg/dL, there was a significant correlation between mean values of D-LDL-C and C-LDL-C (r=0.985 and r=0.974, respectively). Compared to other formulas, C-LDL-C obtained from the proposed formula had the highest correlation with D-LDL-C. The agreement between D-LDL-C and C-LDL-C for TC<200, 200-239, and ≥240 mg/dL was 80.8%, 63.2%, and 67.4%, respectively, indicating a higher level of agreement than other formulas. Conclusion: The new formula appears to be more accurate than FF when applied to the population of Southern Iran.

Novel Probiotic Bacterium Rouxiella badensis subsp. acadiensis (Canan SV-53) Modulates Gut Immunity through Epigenetic Mechanisms.

Gut immune system homeostasis is crucial to overall host health. Immune disturbance at the gut level may lead to systemic and distant sites' immune dysfunction. Probiotics and prebiotics consumption have been shown to improve gut microbiota composition and function and enhance gut immunity. In the current study, the immunomodulatory and anti-inflammatory effects of viable and heat-inactivated forms of the novel probiotic bacterium Rouxiella badensis subsp. acadiensis (Canan SV-53), as well as the prebiotic protocatechuic acid (PCA) derived from the fermentation of blueberry juice by SV-53, were examined. To this end, female Balb/c mice received probiotic (viable or heat-inactivated), prebiotic, or a mixture of viable probiotic and prebiotic in drinking water for three weeks. To better decipher the immunomodulatory effects of biotics intake, gut microbiota, gut mucosal immunity, T helper-17 (Th17) cell-related cytokines, and epigenetic modulation of Th17 cells were studied. In mice receiving viable SV-53 and PCA, a significant increase was noted in serum IgA levels and the number of IgA-producing B cells in the ileum. A significant reduction was observed in the concentrations of proinflammatory cytokines, including interleukin (IL)-17A, IL-6, and IL-23, and expression of two proinflammatory miRNAs, miR-223 and miR425, in treated groups. In addition, heat-inactivated SV-53 exerted immunomodulatory properties by elevating the IgA concentration in the serum and reducing IL-6 and IL-23 levels in the ileum. DNA methylation analysis revealed the role of heat-inactivated SV-53 in the epigenetic regulation of genes related to Th17 and IL-17 production and function, including Il6, Il17rc, Il9, Il11, Akt1, Ikbkg, Sgk1, Cblb, and Smad4. Taken together, these findings may reflect the potential role of the novel probiotic bacterium SV-53 and prebiotic PCA in improving gut immunity and homeostasis. Further studies are required to ascertain the beneficial effects of this novel bacterium in the inflammatory state.

The effect of topical olive oil dressing on the healing of grade 1 and 2 diabetic foot ulcers: An assessor-blind randomized controlled trial in type 2 diabetes patients.

BACKGROUND AND AIMS: We aimed to compare the effect of topical olive oil dressing plus standard care with standard care alone on the treatment of grade 1 and 2 diabetic foot ulcers (DFUs) in type 2 diabetes mellitus (T2DM) patients. METHODS: This assessor-blind randomized controlled trial included 60 T2DM patients with DFU referred to the Diabetes Clinic of Shahid Mohammadi Hospital, Bandar Abbas, Iran, from February 21 to August 22, 2017. Patients were randomly assigned to intervention (n = 30) and control (n = 30) groups. The intervention group received standard care, including wound irrigation with normal saline and oral antibiotics plus daily topical olive oil dressing for four weeks, and the control group only received standard care. The wound healing assessment scale (wound degree, color, drainage, and surrounding tissue healing) was recorded weekly and the total wound status was determined at the end of the study. RESULTS: Treatment with olive oil led to significantly higher scores of ulcer degree, color, drainage, and surrounding tissue healing at weeks one, two, three and four in the olive oil group than in the control group (P < 0.001). Also, the total wound status score was higher in the olive oil group compared to the control group (P < 0.001). The proportions of completely healed, partially healed, and unhealed wounds were 76.6%, 23.3% and 0% in the intervention group, and 0%, 93.3% and 6.7% in the control group, respectively. CONCLUSIONS: Topical olive oil dressing promoted the healing of DFU and it can be recommended as a safe and effective treatment in this regard. TRIAL REGISTRATION: Iranian Registry of Clinical Trials (IRCT), IRCT20150607022585N4. Registered 05/12/2018. Retrospectively registered, https://www.irct.ir/trial/19460.

Pre-diabetes, Diabetes Mellitus and Related Cardio-metabolic Risk Factors in the Southern Coastal Region of Iran Middle-aged and Elderly Population; Bandare-Kong Cohort Study.

BACKGROUND: To evaluate the prevalence of type 2 diabetes mellitus (T2DM), impaired fasting glucose (IFG), and its cardio-metabolic risk factors in the southern Iranian adult population. METHODS: This is a population-based cross-sectional survey on 3944 middle-aged and elderly adults (35-70 years) from Bandare-Kong. The participants were recruited from 2016 to 2018 and the first phase data of the Bandare-Kong Cohort as a part of the PERSIAN Cohort were used for analysis. RESULTS: Among the 3944 included adults, the age-adjusted prevalence of T2DM and IFG was 17.40% and 20.61%, respectively. Mean FPG was higher among those older than 55 years, females, rural residents, current cigarette smokers, hypertriglyceridemia, hypercholesterolemia, unemployed and low educational level in subjects with diabetes and pre-diabetes. T2DM and IFG were more prevalent in women and men, respectively. Also, those with higher waist circumference (WC), higher body mass index (BMI), lower educational levels, rural residents, former cigarette smokers, hypertension (HTN), hypercholesterolemia, hypertriglyceridemia and age older 45 years, had a higher T2DM and IFG prevalence. Multivariable regression analysis showed that older age, higher WC, HTN and hypertriglyceridemia and living in rural regions were statistically significant predictors of T2DM and pre-diabetes while BMI≥25 kg/m2 was the only significant risk factor for IFG. CONCLUSION: The current study illustrated that T2DM and IFG have a high prevalence among the middle-aged and elderly adult Iranian population, particularly in rural dwellers. Hence, prevention strategies should be implemented to reduce diabetes and pre-diabetes, especially in rural areas.

Anti-Inflammatory and Immunomodulatory Properties of Fermented Plant Foods.

Fermented plant foods are gaining wide interest worldwide as healthy foods due to their unique sensory features and their health-promoting potentials, such as antiobesity, antidiabetic, antihypertensive, and anticarcinogenic activities. Many fermented foods are a rich source of nutrients, phytochemicals, bioactive compounds, and probiotic microbes. The excellent biological activities of these functional foods, such as anti-inflammatory and immunomodulatory functions, are widely attributable to their high antioxidant content and lactic acid-producing bacteria (LAB). LAB contribute to the maintenance of a healthy gut microbiota composition and improvement of local and systemic immunity. Besides, antioxidant compounds are involved in several functional properties of fermented plant products by neutralizing free radicals, regulating antioxidant enzyme activities, reducing oxidative stress, ameliorating inflammatory responses, and enhancing immune system performance. Therefore, these products may protect against chronic inflammatory diseases, which are known as the leading cause of mortality worldwide. Given that a large body of evidence supports the role of fermented plant foods in health promotion and disease prevention, we aim to discuss the potential anti-inflammatory and immunomodulatory properties of selected fermented plant foods, including berries, cabbage, and soybean products, and their effects on gut microbiota.

A Pediatric Parathyroid Carcinoma: An Unusual Clinical Presentation and Mini-review.

INTRODUCTION: Primary hyperparathyroidism (PHPT) is a rare condition in the pediatric population. Parathyroid carcinoma (PC) is a very uncommon cause of PHPT, accounting for < 1% of pediatric PHPT cases. It is challenging to distinguish between parathyroid adenoma (PA), the most common cause of PHPT, and PC. In this report, we described a young female who presented with a history of progressive limping and was finally diagnosed with PC. CASE PRESENTATION: A 15-year-old girl presented with progressive limping and bone pain for 8 years. She was referred by an orthopedic surgeon because of elevated intact parathyroid hormone (iPTH) for further evaluation. Physical examination revealed a large, firm, and non-tender neck mass, left hip tenderness, and limited range of motion. The initial biochemistry tests showed a borderline high calcium level of 10.8 mg/dl, an elevated iPTH level of 2876 pg/mL, and a decreased phosphorus level of 2.4 mg/dL. The 99mTechnetium (Tc) sestamibi scan displayed early intense activity in the right thyroid lobe persisting in the three-hour repeat scan, compatible with a parathyroid lesion. The patient underwent right-sided neck exploration and parathyroidectomy. Intraoperative and pathology findings confirmed the diagnosis of PC. Immunohistochemistry (IHC) staining revealed creatine kinase (CK) and CD31 in endothelial cells of the tumor. Ki67 staining was also positive in 2% - 3% of tumor cells. The whole exome sequencing (WES) study was negative for cell division cycle 73 (CDC73) and multiple endocrine neoplasia 1 (MEN1) genes. CONCLUSIONS: PC should be considered as a differential diagnosis of PHPT in the pediatric population, even in the presence of mild hypercalcemia.

Hesperidin inhibits insulin-induced phosphoinositide 3-kinase/Akt activation in human pre-B cell line NALM-6.

CONTEXT: It has been shown that hesperidin induces apoptosis in NALM-6 cells through inhibition of nuclear factor-kappa B (NF-κB) activation. AIMS: To investigate the effect of hesperidin on inhibition of NF-κB activation through blocking phosphoinositide 3-kinase (PI3K)/Akt pathway as a main target in cancer treatment, in NALM-6 cells. MATERIALS AND METHODS: NALM-6 cells were incubated with two concentrations of hesperidin (25, 50 µM) in the presence or absence of insulin (100 nM), as a potent activator of Akt. The cytotoxic activity of hesperidin was determined by 3-(4,5-methylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Cell apoptotic death was measured by ELISA test using cell death detection ELISA Plus kit. To assay the effect of hesperidin on Akt pathway, the phosphorylation levels of Akt, inhibitor of kappa B alpha (IκBα), and glycogen synthase kinase-3 beta (GSK-3ß) and expression level of IκB kinase alpha (IKKα) were determined by Western blot analysis. RESULTS: Hesperidin (both concentrations) significantly reduced cells survival in the presence and absence of insulin compared to untreated cells in a time-dependent manner (P < 0.05). Hesperidin also significantly increased apoptosis in NALM-6 cells even in hyperinsulinemia condition (P < 0.0001). Hesperidin inhibited insulin-induced phosphorylation and activation of Akt, IκBα, and GSK-3ß and decreased expression of IKKα. CONCLUSION: The results of this study demonstrated that cytotoxic and proapoptotic actions of hesperidin are partly mediated through the suppression of PI3K3/Akt/IKK signaling pathway. So, hesperidin might act as a chemotherapeutic agent by targeting cell survival pathways.

High Body Mass Index and Young Age Are not Associated with Post-Mastectomy Pain Syndrome in Breast Cancer Survivors: A Case-Control Study.

BACKGROUND: Surgery is usually the first treatment for breast cancer which is followed by some complications such as chronic pain. Post mastectomy pain syndrome (PMPS) is a common complication among breast cancer survivors and is considered as a chronic neuropathic pain in the side of surgery which persists more than three months. The exact mechanisms and related risk factors of the chronic pain after breast surgery are unknown. The aim of this study was to investigate the association of body mass index (BMI) and age with PMPS. METHODS: In this case-control study, a total of 122 women were assessed; of these, 61 women were diagnosed with PMPS and selected as cases and 61 pain-free patients were selected as controls. The demographic and clinical characteristics of participants were collected through questionnaires and medical record of patients. Logistic regression model was used to determine the association of BMI and age with PMPS, adjusted for demographic and clinical characteristics. RESULTS: No significant differences were found in means of weight (68.02±8.80 vs. 68.67±11.82, p=0.726), BMI (26.38±3.28 vs. 27.10±6.03, p=0.410), and age (46.34±11.67 vs. 48.54±12.57, p=0.319) between those with PMPS and those not reporting PMPS. A non-significant slight increase in odds ratio of PMPS was observed in obese category compared to normal weight category [OR=1.152 (95% CI 0.405-3.275), p=0.908], but after adjusting the confounding factors, the risk of pain development was attenuated in obese subjects [OR=0.748 (95% CI 0.228-2.459), p=0.633]. Also, non-significant decrease in odds ratios of PMPS was found in 20-39 y, 40-49 y, and 50-59 y ages categories compared to oldest age category [adjusted OR= 0.781 (95% CI 0.213-2.866), p=0.576; adjusted OR=0.485 (95% CI 0.152- 1.554), p=0.183; adjusted OR=0.735 (95% CI 0.206-2.627), p=0.628; respectively]. CONCLUSION: In contrast with some observational studies, present study showed that high BMI and younger age might not be associated with increased risk of PMPS development. Further research is necessary to determine the main risk factors and directionality and causal mechanisms for associations of these risk factors with chronic pain after mastectomy.