Attenuation of brown adipocyte whitening in high-fat diet-induced obese rats: Effects of melatonin and ß-hydroxybutyrate on Cidea, Fsp27 and MT1 expression.

AIM: To investigate the effects of ß-hydroxybutyrate (BHB) and melatonin on brown adipose tissue (BAT) plasticity in rats fed a high-fat diet (HFD). METHODS: We employed a 7-week experimental design for a study on 30 male Sprague-Dawley rats divided into five groups: (1) a control-diet fed group; (2) a high-fat diet (HFD)-fed group; (3) a group that received an HFD and a BHB solution in their drinking water; (4) a group that received an HFD with 10 mg/kg/day melatonin in their drinking water; and (5) a group that received an HFD and were also treated with the combination of BHB and melatonin. Following the treatment period, biochemical indices, gene expression levels of key thermogenic markers (including uncoupling protein 1 [UCP1], PR domain containing 16 [PRDM16], Cidea, fat-specific protein 27 [Fsp27], and metallothionein 1 [MT1]), and stereological assessments of BAT were evaluated. RESULTS: Treatment with BHB and melatonin significantly boosted blood ketone levels, improved lipid profiles, and reduced weight gain from an HFD. It also downregulated genes linked to WAT, namely, Cidea and Fsp27, and upregulated key BAT markers, including UCP1, PRDM16 and peroxisome proliferator-activated receptor-gamma coactivator-1-alpha. Additionally, the co-treatment increased MT1 receptor expression and enhanced the structural density of BAT. CONCLUSION: The combined oral administration of BHB and melatonin successfully prevented the whitening of BAT in obese rats fed an HFD, indicating its potential as a therapeutic strategy for obesity-related BAT dysfunction. The synergistic effects of this treatment underscore the potential of a combined approach to address BAT dysfunction in obesity.

The effects of metformin and PCL-sorafenib nanoparticle co-treatment on MCF-7 cell culture model of breast cancer.

Despite breakthrough therapeutics in breast cancer, it is one of the main causes of mortality among women worldwide. Thus, drug therapies for treating breast cancer have recently been developed by scientists. Metformin and sorafenib are well-known therapeutics in breast cancer. In the present study, we combined sorafenib and PCL-sorafenib with metformin to improve drug absorption and promote therapeutic efficiency. The MCF-7 cells were treated with metformin, sorafenib, or PCL-sorafenib. The growth inhibitory effect of these drugs and cell viability were assessed using MTT and flow cytometry assays, respectively. The expression of targeted genes involved in cell proliferation, signaling, and the cell cycle was measured by real-time PCR. The results showed that MCF-7 cells treated with metformin/sorafenib and PCL-sorafenib/metformin co-treatment contributed to 50% viability compared to the untreated group. Moreover, PI and Annexin V staining tests showed that the cell viability for metformin/sorafenib and PCL-sorafenib/metformin was 38% and 17%, respectively. Furthermore, sorafenib/metformin and PCL-sorafenib/metformin lead to p53 gene expression increase by which they can increase ROS, thereby decreasing GPX4 gene expression. In addition, they affected the expression of BCL2 and BAX genes and altered the cell cycle. Together, the combination of PCL-sorafenib/metformin and sorafenib/metformin increased sorafenib absorption at lower doses and also led to apoptosis and oxidative stress increases in MCF-7 cells.

Bilirubin, an endogenous antioxidant that affects p53 protein and its downstream apoptosis/autophagy-related genes in LS180 and SW480 cell culture models of colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is one of the commonest neoplasms worldwide, which its pathogenesis is strongly correlated with p53 mutations. Antioxidants are believed to decelerate the CRC progression, possibly through interfering with p53 and its downstream target genes and mechanisms. Regarding the potential antioxidant effects of bilirubin, as an incredible endogenous antioxidant, we sought to investigate how bilirubin affected the expression levels of p53 protein and its downstream target genes, including Mdm2, Bcl-2, BECN1 and LC3, in LS180 and SW480 cell culture models of CRC. METHODS AND RESULTS: Using the MTT (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-2H-tetrazolium bromide) assay, 50 and 100 µM concentrations of bilirubin were determined to be non-toxic for both LS180 and SW480 cell lines. Western blot analysis was employed to evaluate the protein expression levels of p53. The results revealed that p53 protein levels were higher in LS180 cells treated with bilirubin compared to the control group. Notwithstanding, in SW480 cells, no considerable changes were observed in p53 protein levels of treated cells compared to the control ones. The quantitative reverse transcriptase-polymerase chain reaction (q RT-PCR) method was used to measure the mRNA expression levels of the apoptosis/autophagy-related genes, Mdm2, Bcl-2, BECN1, and LC3 , as the p53's downstream target genes. Consequently, the expression of Bcl-2 and Mdm2 genes were affected by p53, while BECN1 and LC3 expression levels were decreased in both cell lines. CONCLUSION: Bilirubin is an endogenous antioxidant with significant anti-tumor effects in the studied CRC cell lines, probably through the regulation of p53 protein expression levels and subsequent control of apoptosis and autophagy, as two key processes involved in cell survival and progression of tumor cells.

The crosstalk between ubiquitin-conjugating enzyme E2Q1 and p53 in colorectal cancer: An in vitro analysis.

Colorectal cancer (CRC) is a prevalent gastrointestinal neoplasm that ranks fourth in terms of cancer-related deaths worldwide. In the process of CRC progression, multiple ubiquitin-conjugating enzymes (E2s) are involved; UBE2Q1 is one of those newly identified E2s that is markedly expressed in human colorectal tumors. Since p53 is a well-known tumor suppressor and defined as a key factor to be targeted by the ubiquitin-proteasome system, we hypothesized that UBE2Q1 might contribute to CRC progression through the modulation of p53. Using the lipofection method, the cultured SW480 and LS180 cells were transfected with the UBE2Q1 ORF-containing pCMV6-AN-GFP vector. Then, quantitative RT-PCR was used to assay the mRNA expression levels of p53's target genes, i.e., Mdm2, Bcl2, and Cyclin E. Moreover, Western blot analysis was performed to confirm the cellular overexpression of UBE2Q1 and assess the protein levels of p53, pre- and post-transfection. The expression of p53's target genes were cell line-dependent except for Mdm2 that was consistent with the findings of p53. The results of Western blotting demonstrated that the protein levels of p53 were greatly lower in UBE2Q1-transfected SW480 cells compared to the control SW480 cells. However, the reduced levels of p53 protein were not remarkable in the transfected LS180 cells compared to the control cells. The suppression of p53 is believed to be the result of UBE2Q1-dependent ubiquitination and its subsequent proteasomal degradation. Furthermore, the ubiquitination of p53 can act as a signal for degradation-independent functions, such as nuclear export and suppressing the p53's transcriptional activities. In this context, the decreased Mdm2 levels can moderate the proteasome-independent mono-ubiquitination of p53. The ubiquitinated p53 modulates the transcriptional levels of target genes. Therefore, the up-modulation of UBE2Q1 may influence the transcriptional activities depending on p53, and thereby contributes to CRC progression through regulating the p53.

In Vitro and In Vivo Improvement of Islet Quality and Transplantation Successes following Islet Treatment with Biomaterials in Diabetic Rats.

Background: Loss of islet survival and function, caused by native niche disruption and oxidative stress induction during mechanical and enzymatic isolation, limits the effectiveness of islet transplantation. Reconstitution of islet microenvironment, vascularization, and decreased oxidative stress with biomaterials may improve islet quality and graft outcomes. We investigated effects of two biomaterials, platelet-rich plasma and pancreatic islets homogenate combination on islet recovery and quality by evaluating in vitro islet survival, secretory function, and oxidative stress parameters and assessing in vivo transplantation outcomes. Methods: In vitro, islet viability and secretory function of isolated islets were assessed after 24 h and 72 h incubation with biomaterials. Also, oxidative stress markers were measured once after isolation and 24 h after incubation with biomaterials. For evaluating in vivo effects, cultured islets for 24 h were transplanted into subscapular space of diabetic rat kidney, and outcomes were analyzed by measuring serum glucose and insulin concentrations, glucose tolerance test, level of oxidative parameters, and pancreatic gene expression. Results: Treating islets with biomaterials significantly increased their viability and secretory function, reduced MDA level, and elevate SOD and CAT activity. Decreased level of glucose and MDA improved insulin level, increased SOD activity, and also enhanced pdx1 and insulin gene expression in diabetic rats after islet transplantation. Conclusions: Biomaterials used in the present study should be consider as beneficial materials for increasing islet transplantation outcome. These materials may hamper transplantation limitation to some extent.