Category Archives: BioFactors

Rat brown adipose tissue thermogenic markers are modulated by estrous cycle phases and short‐term fasting

Posted on 21 July 2023 by

This study confirmed that BAT exhibits morphological and functional changes in proestrus and diestrus. Moreover, BAT undergoes additional dynamic functional and morphological changes during short-term fasting.

Abstract

Brown adipose tissue (BAT) converts chemical energy into heat to maintain body temperature. Although fatty acids (FAs) represent a primary substrate for uncoupling protein 1 (UCP1)-dependent thermogenesis, BAT also utilizes glucose for the same purpose. Considering that estrous cycle effects on BAT are not greatly explored, we examined those of 6-h fasting on interscapular BAT (iBAT) thermogenic markers in proestrus and diestrus. We found that the percentage of multilocular adipocytes was lower in proestrus than in diestrus, although it was increased after fasting in both analyzed estrous cycle stages. Furthermore, the percentage of paucilocular adipocytes was increased by fasting, unlike the percentage of unilocular cells, which decreased in both analyzed stages of the estrous cycle. The UCP1 amount was lower in proestrus irrespectively of the examined dietary regimens. Regarding FA transporters, it was shown that iBAT CD36 content was increased in fasted rats in diestrus. In contrast to GLUT1, the level of GLUT4 was interactively modulated by selected estrous cycle phases and fasting. There was no change in insulin receptor and ERK1/2 activation, while AKT activation was interactively modulated by fasting and estrous cycle stages. Our study showed that iBAT exhibits morphological and functional changes in proestrus and diestrus. Moreover, iBAT undergoes additional dynamic functional and morphological changes during short-term fasting to modulate nutrient utilization and adjust energy expenditure.

Protein concentrations and activities of fatty acid desaturase and elongase enzymes in liver, brain, testicle, and kidney from mice: Substrate dependency

Posted on 20 July 2023 by

The liver had the highest capacity for PUFA biosynthesis, with limited activity in the brain, testicles, and kidney, while we failed to detect activity in the heart and lung. The protein content and activity of the enzymes were significantly correlated. The capacity for PUFA synthesis in mice mainly resides in the liver, with enzymes having preference for n-3 PUFAs.

Abstract

The synthesis rates of n-3 and n-6 polyunsaturated fatty acids (PUFAs) in rodents and humans are not agreed upon and depend on substrate availability independently of the capacity for synthesis. Therefore, we aimed to assess the activities of the enzymes for n-3 and n-6 PUFA synthesis pathways in liver, brain, testicle, kidney, heart, and lung, in relation to their protein concentration levels. Eight-week-old Balb/c mice (n = 8) were fed a standard chow diet (6.2% fat, 18.6% protein, and 44.2% carbohydrates) until 14 weeks of age, anesthetized with isoflurane and tissue samples were collected (previously perfused) and stored at −80°C. The protein concentration of the enzymes (Δ-6D, Δ-5D, Elovl2, and Elovl5) were assessed by ELISA kits; their activities were assayed using specific PUFA precursors and measuring the respective PUFA products as fatty acid methyl esters by gas chromatographic analysis. The liver had the highest capacity for PUFA biosynthesis, with limited activity in the brain, testicles, and kidney, while we failed to detect activity in the heart and lung. The protein concentration and activity of the enzymes were significantly correlated. Furthermore, Δ-6D, Δ-5D, and Elovl2 have a higher affinity for n-3 PUFA precursors compared to n-6 PUFA. The capacity for PUFA synthesis in mice mainly resides in the liver, with enzymes having preference for n-3 PUFAs.

Sodium Danshensu ameliorates cerebral ischemia/reperfusion injury by inhibiting CLIC4/NLRP3 inflammasome‐mediated endothelial cell pyroptosis

Posted on 17 July 2023 by

Mechanism of SDSS in inhibiting endothelial cell pyroptosis. In the priming step, NLRP3, pro-Caspase-1, GSDMD-full, pro-IL-1β, and pro-IL-18 were up-regulated. Furthermore, the translocation of CLIC4 from cytoplasm to the membrane induced chloride outflow, resulting in the assembly of NLRP3, ASC and Pro-Caspase-1 into a platform (activation step). By binding CLIC4 and blocking its membrane localization, SDSS inhibited chloride outflow, thus inhibiting the activation of NLRP3 inflammasome and then the cleavage of pro-Caspase-1 into Caspase-1. This inhibited pyroptosis along with the release of IL-1β and IL-18, resulting from Caspase-1-dependent GSDMD-N cleavage.

Abstract

Endothelial pyroptosis promotes cerebral ischemia/reperfusion injury (CIRI). Sodium Danshensu (SDSS) has been shown to attenuate CIRI and have anti-inflammatory properties in endothelial cells. However, the mechanism and effect of SDSS on alleviating endothelial pyroptosis after CIRI remains poorly understood. Thus, we aimed to investigate the efficacy and mechanism of SDSS in reducing endothelial pyroptosis. It has been shown that SDSS administration inhibited NLRP3 inflammasome-mediated pyroptosis. As demonstrated by protein microarrays, molecular docking, CETSA and ITDRF_CETSA, SDSS bound strongly to CLIC4. Furthermore, SDSS can decrease its expression and inhibit its translocation. Its effectiveness was lowered by CLIC4 overexpression but not by knockdown. Overall The beneficial effect of SDSS against CIRI in this study can be ascribed to blocking endothelial pyroptosis by binding to CLIC4 and then inhibiting chloride efflux-dependent NLRP3 inflammasome activation.

P38α contributes to TNF‐α‐induced IL‐8 production in human gingival cells

Posted on 14 July 2023 by nDan Mao, nHiroshi Inoue, nTakuya Notomi, nSeiji Godan

When phosphorylated at threonine 180 and tyrosine 182, p38α contributes to the induction of IL-8 by TNF-α in Ca9-22 cells. TNF-α-induced phosphorylation translocates NF-κB into the nucleus and then increases both IL-8 mRNA expression and secretion. Thus, p38α plays an important role in TNF-α-induced IL-8 production, providing a potential therapeutic target to prevent and treat periodontal disease.

Abstract

Tumor necrosis factor-alpha (TNF-α) is a major inflammatory cytokine that induces interleukin (IL)-8 production. Although some studies have reported the involvement of the p38 MAPK signaling pathway in TNF-α-induced IL-8 production, its specific regulatory mechanisms in gingival epithelial cells (GECs) are still poorly understood. In the present study, Ca9-22 cells were used as representative GECs to investigate the effect of p38 signaling on TNF-α-induced IL-8 production. We found that TNF-α enhanced IL-8 production in Ca9-22 cells by activating the p38 signaling pathway, and one of its isoforms, p38α, played a key role. P38α deletion markedly inhibited TNF-α-induced IL-8 expression in Ca9-22 cells, while p38α gene rescue could reverse this effect. Further studies revealed that TNF-α-induced IL-8 production was markedly reduced when the threonine 180 and tyrosine 182 p38α phosphorylation sites were targeted for mutagenesis to alanine and phenylalanine, respectively, suggesting their critical role in the process. In conclusion, p38α plays an important role in TNF-α-induced IL-8 production, providing a potential therapeutic target to prevent and treat periodontal disease.

A novel lncRNA LOC105613571 binding with BDNF in pituitary promotes gonadotropin secretion by AKT/ERK‐mTOR pathway in sheep associated with prolificacy

Posted on 11 July 2023 by

Candidate lncRNA LOC105613571 targeted BDNF via trans-regulated relationships was characterized by pituitary transcriptome from Hu sheep with high and low fecundity. GnRH stimulation increased BDNF and lncRNA LOC105613571 expression in pituitary cells. BDNF-binding lncRNA LOC105613571 promotes pituitary gonadotropin secretion by activating AKT/ERK-mTOR pathway in pituitary cells.

Abstract

The pituitary is a vital endocrine organ for synthesis and secretion of gonadotropic hormones (FSH and LH), and the gonadotropin showed fluctuations in animals with different fecundity. Long non-coding RNAs (lncRNAs) have been identified as regulatory factors for the reproductive process. However, the profiles of lncRNAs and their roles involved in sheep fecundity remains unclear. In this study, we performed RNA-sequencing for the sheep pituitary gland associated with different fecundity, and identified a novel candidate lncRNA LOC105613571 targeting BDNF related to gonadotropin secretion. Our results showed that expression of lncRNA LOC105613571 and BDNF could be significantly upregulated by GnRH stimulation in sheep pituitary cells in vitro. Notably, either lncRNA LOC105613571 or BDNF silencing inhibited cell proliferation while promoted cell apoptosis. Moreover, lncRNA LOC105613571 knockdown could also downregulate gonadotropin secretion via inactivation AKT, ERK and mTOR pathway. In addition, co-treatment with GnRH stimulation and lncRNA LOC105613571 or BDNF knockdown showed the opposite effect on sheep pituitary cells in vitro. In summary, BDNF-binding lncRNA LOC105613571 in sheep regulates pituitary cell proliferation and gonadotropin secretion via the AKT/ERK-mTOR pathway, providing new ideas for the molecular mechanisms of pituitary functions.

The influence of redox modulation on hypoxic endothelial cell metabolic and proteomic profiles through a small thiol‐based compound tuning glutathione and thioredoxin systems

Posted on 6 July 2023 by

The intracellular redox state of endothelial cells facing low oxygen and oxidative stress was regulated via the pro-glutathione molecule I-152, a co-drug of N-acetylcysteine and cysteamine. The principal redox couples, GSH/GSSG, and NAD(P)+/NAD(P)H were affected by hypoxia and in turn, modulated with I-152. Glutathione and thioredoxin-related pathways were enhanced after treatment and ROS production was alleviated. Strategies to fine-tune the redox balance could ameliorate the cell response to hypoxic environments.

Abstract

Reduction in oxygen levels is a key feature in the physiology of the bone marrow (BM) niche where hematopoiesis occurs. The BM niche is a highly vascularized tissue and endothelial cells (ECs) support and regulate blood cell formation from hematopoietic stem cells (HSCs). While in vivo studies are limited, ECs when cultured in vitro at low O₂ (<5%), fail to support functional HSC maintenance due to oxidative environment. Therefore, changes in EC redox status induced by antioxidant molecules may lead to alterations in the cellular response to hypoxia likely favoring HSC self-renewal. To evaluate the impact of redox regulation, HUVEC, exposed for 1, 6, and 24 h to 3% O₂ were treated with N-(N-acetyl-l-cysteinyl)-S-acetylcysteamine (I-152). Metabolomic analyses revealed that I-152 increased glutathione levels and influenced the metabolic profiles interconnected with the glutathione system and the redox couples NAD(P)+/NAD(P)H. mRNA analysis showed a lowered gene expression of HIF-1α and VEGF following I-152 treatment whereas TRX 1 and 2 were stimulated. Accordingly, the proteomic study revealed the redox-dependent upregulation of thioredoxin and peroxiredoxins that, together with the glutathione system, are the main regulators of intracellular ROS. Indeed, a time-dependent ROS production under hypoxia and a quenching effect of the molecule were evidenced. At the secretome level, the molecule downregulated IL-6, MCP-1, and PDGF-bb. These results suggest that redox modulation by I-152 reduces oxidative stress and ROS level in hypoxic ECs and may be a strategy to fine-tune the environment of an in vitro BM niche able to support functional HSC maintenance.

Nomilin and its analogue obacunone alleviate NASH and hepatic fibrosis in mice via enhancing antioxidant and anti‐inflammation capacity

Posted on 4 July 2023 by

Nomilin and obacunone exert beneficial effects on MCD-, BDL-, and CCl₄-induced NASH mice. Nomilin and obacunone alleviate NASH and liver fibrosis via enhancing antioxidant and anti-inflammation capacity.

Abstract

Nonalcoholic steatohepatitis (NASH) and hepatic fibrosis are leading causes of cirrhosis with rising morbidity and mortality worldwide. Currently, there is no appropriate treatment for NASH and hepatic fibrosis. Many studies have shown that oxidative stress is a main factor inducing NASH. Nomilin (NML) and obacunone (OBA) are limonoid compounds naturally occurring in citrus fruits with various biological properties. However, whether OBA and NML have beneficial effects on NASH remains unclear. Here, we demonstrated that OBA and NML inhibited hepatic tissue necrosis, inflammatory infiltration and liver fibrosis progression in methionine and choline-deficient (MCD) diet, carbon tetrachloride (CCl₄)-treated and bile duct ligation (BDL) NASH and hepatic fibrosis mouse models. Mechanistic studies showed that NML and OBA enhanced anti-oxidative effects, including reduction of malondialdehyde (MDA) level, increase of catalase (CAT) activity and the gene expression of glutathione S-transferases (GSTs) and Nrf2-keap1 signaling. Additional, NML and OBA inhibited the expression of inflammatory gene interleukin 6 (Il-6), and regulated the bile acid metabolism genes Cyp3a11, Cyp7a1, multidrug resistance-associated protein 3 (Mrp3). Overall, these findings indicate that NML and OBA may alleviate NASH and liver fibrosis in mice via enhancing antioxidant and anti-inflammation capacity. Our study proposed that NML and OBA may be potential strategies for NASH treatment.

Correction to “Inhibition of succinate dehydrogenase sensitizes cyclin E‐driven ovarian cancer to CDK inhibition”

Posted on 30 June 2023 by Wiley: BioFactors: Table of Contents

BioFactors, EarlyView.

Rigosertib is more potent than wortmannin and rapamycin against adult T‐cell leukemia‐lymphoma

Posted on 22 June 2023 by

HTLV-1 downregulation of the mRNA level may occur as a negative feedback response to increased PI3K-Akt-mTOR phosphorylation by HTLV-1. Rigosertib was more effective than wortmannin and rapamycin in inducing cell cycle arrest, as well as a significant late apoptosis in the Inf-3T3 and MT-2 cells.

Abstract

Human T lymphotropic virus type 1 (HTLV-1) infection can cause adult T-cell lymphoblastic leukemia (ATLL), an incurable, chemotherapy-resistant malignancy. In a quest for new therapeutic targets, our study sought to determine the levels of AKT, mTOR, and PI3K in ATLL MT-2 cells, HTLV-1 infected NIH/3T3 cells (Inf-3T3), and HTLV-1 infected patients (Carrier, HAM/TSP, and ATLL). Furthermore, the effects of rigosertib, wortmannin, and rapamycin on the PI3K/Akt/mTOR pathway to inhibit the proliferation of ATLL cells were examined. The results showed that mRNA expression of Akt/PI3K/mTOR was down-regulated in carrier, HAM/TSP, and ATLL patients, as well as MT-2, and Inf-3T3 cells, compared to the healthy individuals and untreated MT-2 and Inf-3T3 as controls. However, western blotting revealed an increase in the phosphorylated and activated forms of AKT and mTOR. Treating the cells with rapamycin, wortmannin, and rigosertib decreased the phosphorylated forms of Akt and mTOR and restored their mRNA expression levels. Using these inhibitors also significantly boosted the expression of the pro-apoptotic genes, Bax/Bcl-2 ratio as well as the expression of the tumor suppressor gene p53 in the MT-2 and Inf-3T3cells. Rigosertib was more potent than wortmannin and rapamycin in inducing sub-G1 and G2-M cell cycle arrest, as well as late apoptosis in the Inf-3T3 and MT-2 cells. It also synergized the cytotoxic effects of vincristine. These findings demonstrate that HTLV-1 downregulation of the mRNA level may occur as a negative feedback response to increased PI3K-Akt-mTOR phosphorylation by HTLV-1. Therefore, using rigosertib alone or in combination with common chemotherapy drugs may be beneficial in ATLL patients.

SUMO2/3 promotes the progression and oxaliplatin resistance of colorectal cancer through facilitating the SUMOylation at Ku80‐K307

Posted on 20 June 2023 by

We describes the role for SUMO2/3 during oxaliplatin resistance in CRC and assessed the contribution of Ku80 SUMOylation in this process. Overexpression of SUMO2/3-promoted CRC cell proliferation, invasion, migration in vitro and in vivo. SUMOylation of K307 in Ku80 attenuated the DNA damage of CRC cells caused by oxaliplatin and was consistent with an antiapoptotic role for SUMO2/3.

Abstract

Colorectal cancer (CRC) is one of the most prevalent cancers worldwide and is typically treated with the FOLFOX regimen (folinic acid, 5-fluorouracil, and oxaliplatin). However, oxaliplatin resistance remains a serious clinical problem. In the present study, we found that SUMO2/3 was overexpressed in CRC tissues and exogenous overexpression of SUMO2/3 promoted CRC cell proliferation, extension, and invasion and positively regulated the cell cycle. In contrast, SUMO2/3 gene knockdowns inhibited migration and repressed cell viability in vitro and in vivo. In addition, we found that SUMO2/3 was recruited to the cell nucleus and suppressed oxaliplatin-induced apoptosis of CRC cells. Moreover, Ku80, a DNA-binding protein essential for the repair of DNA double-strand breaks, was confirmed to bind with SUMO2/3. Notably, Ku80 undergoes SUMOylation at K307 by SUMO2/3 and this correlated with apoptosis in CRC cells suffering oxaliplatin stress. Collectively, we found that SUMO2/3 plays a specific role in CRC tumorigenesis and acts through Ku80 SUMOylation which is linked with the development of CRC-oxaliplatin resistance.