Uncovering SIRT3 and SHMT2-dependent pathways as novel targets for apigenin in modulating colorectal cancer: In vitro and in vivo studies.

Despite significant advances in the treatment of colorectal cancer (CRC), identification of novel targets and treatment options are imperative for improving its prognosis and survival rates. The mitochondrial SIRT3 and SHMT2 have key roles in metabolic reprogramming and cell proliferation. This study investigated the potential use of the natural product apigenin in CRC treatment employing both in vivo and in vitro models and explored the role of SIRT3 and SHMT2 in apigenin-induced CRC apoptosis. The role of SHMT2 in CRC patients' survival was verified using TCGA database. In vivo, apigenin treatment restored the normal colon appearance. On the molecular level, apigenin augmented the immunohistochemical expression of cleaved caspase-3 and attenuated SIRT3 and SHMT2 mRNA expression CRC patients with decreased SHMT2 expression had improved overall and disease-free survival rates. In vitro, apigenin reduced the cell viability in a time-dependent manner, induced G0/G1 cell cycle arrest, and increased the apoptotic cell population compared to the untreated control. Mechanistically, apigenin treatment mitigated the expression of SHMT2, SIRT3, and its upstream long intergenic noncoding RNA LINC01234 in CRC cells. Conclusively, apigenin induces caspase-3-dependent apoptosis in CRC through modulation of SIRT3-triggered mitochondrial pathway suggesting it as a promising therapeutic agent to improve patient outcomes.

Serum lncRNAs TUG1, H19, and NEAT1 and their target miR-29b/SLC3A1 axis as possible biomarkers of preeclampsia: Potential clinical insights.

To date, the epigenetic signature of preeclampsia (PE) is not completely deciphered. Oxidative stress-responsive long non-coding RNAs (lncRNAs) are deregulated in preeclamptic placenta; however, their circulating profiles and diagnostic abilities are still unexplored. We investigated serum redox-sensitive lncRNAs TUG1, H19, and NEAT1, and their target miR-29b/cystine/neutral/dibasic amino acids transporter solute carrier family 3, member 1 (SLC3A1) as potential non-invasive biomarkers of PE risk, onset, and severity. We recruited 82 patients with PE and 78 healthy pregnant women. We classified PE patients into early-onset (EOPE) and late-onset (LOPE) subgroups at a cut-off 34 gestational weeks and into severe and mild PE subgroups by blood pressure and proteinuria criteria. Bioinformatics analysis was employed to select lncRNAs/microRNA/target gene interactions. Serum H19, NEAT1, and SLC3A1 mRNA expression were reduced, meanwhile miR-29b levels were elevated, whereas there was no significant difference in TUG1 levels between PE patients and healthy pregnancies. Serum H19 levels were lower, whereas miR-29b levels were higher in EOPE versus LOPE. Serum miR-29b and H19 levels were higher in severe versus mild PE. ROC analysis identified serum H19, NEAT1, miR-29b, and SLC3A1 as potential diagnostic markers, with H19 (AUC = 0.818, 95%CI = 0.744-0.894) and miR-29b (AUC = 0.82, 95%CI = 0.755-0.885) were superior discriminators. Only H19 and miR-29b discriminated EOPE and severe PE cases. In multivariate logistic analysis, miR-29b and H19 were associated with EOPE, using maternal age and gestational age as covariates, while miR-29b was associated with severe PE, using maternal age as covariate. Studied markers were correlated with clinical and ultrasound data in the overall PE group. Serum H19 and TUG1 were negatively correlated with albuminuria in EOPE and LOPE, respectively. NEAT1 and SLC3A1 were correlated with ultrasound data in EOPE. Likewise, TUG1, miR-29b, and SLC3A1 showed significant correlations with ultrasound data in LOPE. Conclusively, this study configures SLC3A1 expression as a novel potential serum biomarker of PE and advocates serum H19 and miR-29b as biomarkers of EOPE and miR-29b as a biomarker of PE severity.

RNA methylation machinery and m6A target genes as circulating biomarkers of ulcerative colitis and Crohn's disease: Correlation with disease activity, location, and inflammatory cytokines.

Accurate diagnosis of ulcerative colitis (UC) and Crohn's disease (CD), the main subtypes of inflammatory bowel disease (IBD), has been challenging due to the constraints of the current techniques. N6-methyl adenosine (m6A) regulators have evolved as key players in IBD pathogenesis; however, their relation to its clinical setting is largely unexplored. This study investigated the potential of selected RNA methylation machinery and m6A target genes as serum biomarkers of UC and CD, their predictive and discriminating capabilities, and their correlations with laboratory data, interleukin (IL)-6, interferon-γ, disease activity scores, and pathological features. Fifty UC and 45 CD patients, along with 30 healthy volunteers were enlisted. The mRNA expression levels of the m6A writers methyltransferase-like 3 (METTL3) and Wilms-tumor associated protein (WTAP), and the reader YTH domain family, member 1 (YTHDF1), along with the m6A candidate genes sex determining region Y-box 2 (SOX2), hexokinase 2 (HK2), and ubiquitin-conjugating enzyme E2 L3 (UBE2L3) were upregulated in UC patients, whereas only METTL3, HK2, and UBE2L3 were upregulated in CD patients versus controls. Serum WTAP (AUC = 0.94, 95 %CI = 0.874-1.006) and HK2 (AUC = 0.911, 95 %CI = 0.843-0.980) expression levels showed excellent diagnostic accuracy for UC, METTL3 showed excellent diagnostic accuracy for CD (AUC = 0.91, 95 %CI = 0.828-0.992), meanwhile, WTAP showed excellent discriminative power between the two diseases (AUC = 0.91, 95 %CI = 0.849-0.979). Multivariate logistic analysis unveiled the association of METTL3 and UBE2L3 expression with the risk of CD and UC diagnosis, respectively, controlled by age and sex as confounders. Remarkable correlations were recorded between the gene expression of studied m6A regulators and targets in both diseases. Among UC patients, serum METTL3 and WTAP were correlated with UC extent/type, while WTAP was correlated with IL-6. Among CD patients, serum METTL3 and HK2 were correlated with CD activity index (CDAI) and CD location. In conclusion, m6A regulators and target genes are distinctly expressed in UC and CD clinical samples, correlate with disease activity and extent/location, and could serve as a novel approach to empower the diagnosis and stratification of IBD subtypes.

Association of lncRNA MEG3 rs941576 polymorphism, expression profile, and its related targets with the risk of obesity-related colorectal cancer: potential clinical insights.

The identification of novel screening tools is imperative to empower the early detection of colorectal cancer (CRC). The influence of the long non-coding RNA maternally expressed gene 3 (MEG3) rs941576 single nucleotide polymorphism on CRC susceptibility remains uninvestigated. This research appraised MEG3 rs941576 association with the risk and clinical features of CRC and obesity-related CRC and its impact on serum MEG3 expression and its targets miR-27a/insulin-like growth factor 1 (IGF1)/IGF binding protein 3 (IGFBP3) and miR-181a/sirtuin 1 (SIRT1), along with the potential of these markers in obesity-related CRC diagnosis. 130 CRC patients (60 non-obese and 70 obese) and 120 cancer-free controls (64 non-obese and 56 obese) were enrolled. MEG3 targets were selected using bioinformatics analysis. MEG3 rs941576 was associated with magnified CRC risk in overall (OR (95% CI) 4.69(1.51-14.57), P = 0.0018) and stratified age and gender groups, but not with obesity-related CRC risk or MEG3/downstream targets' expression. Escalated miR-27a and IGFBP3 and reduced IGF1 serum levels were concomitant with MEG3 downregulation in overall CRC patients versus controls and obese versus non-obese CRC patients. Serum miR-181a and SIRT1 were upregulated in CRC patients versus controls but weren't altered in the obese versus non-obese comparison. Serum miR-181a and miR-27a were superior in overall and obesity-related CRC diagnosis, respectively; meanwhile, IGF1 was superior in distinguishing obese from non-obese CRC patients. Only serum miR-27a was associated with obesity-related CRC risk in multivariate logistic analysis. Among overall CRC patients, MEG3 rs941576 was associated with lymph node (LN) metastasis and tumor stage, serum MEG3 was negatively correlated with tumor stage, while SIRT1 was correlated with the anatomical site. Significant correlations were recorded between MEG3 and anatomical site, SIRT1 and tumor stage, and miR-27a/IGFBP3 and LN metastasis among obese CRC patients, while IGF1 was correlated with tumor stage and LN metastasis among non-obese CRC patients. Conclusively, this study advocates MEG3 rs941576 as a novel genetic marker of CRC susceptibility and prognosis. Our findings accentuate circulating MEG3/miR-27a/IGF1/IGFBP3, especially miR-27a as valuable markers for the early detection of obesity-related CRC. This axis along with SIRT1 could benefit obesity-related CRC prognosis.

Differential expression of serum miR-486 and miR-25 in ulcerative colitis and Crohn's disease: Correlations with disease activity, extent, and location.

Novel reliable biomarkers of inflammatory bowel disease (IBD) are clinically imperative due to potential limitations of endoscopic techniques. MicroRNAs (miRNAs) have emerged as non-invasive biomarkers of IBD; however, the full disease-specific miRNAs signature for IBD subtypes remains elusive. We evaluated the diagnostic role of circulating miR-486 and miR-25 in IBD patients and their potential ability to discriminate IBD subtypes; ulcerative colitis (UC) and Crohn's disease (CD). Sixty UC patients, 60 CD patients, and 60 healthy controls were recruited. Serum miRNA expression was determined using RT-qPCR. Bioinformatics was employed for target gene and protein-protein interaction (PPI) network analyses. Serum miR-486 was upregulated in CD patients, but didn't change in UC patients compared to controls. Conversely, serum miR-25 was decreased in both CD and UC patients compared to controls. Only miR-486 was differentially expressed between UC and CD patients. Receiver-operating characteristic analysis revealed that serum miR-486 was superior in CD diagnosis (AUC=0.945) and significantly distinguished CD and UC patients, whereas miR-25 showed discriminative potential for both UC and CD from controls. In the multivariate logistic analysis only miR-486 was associated with the risk of CD diagnosis. Serum miR-486 was correlated with CD activity index and location of disease in CD patients, whereas miR-25 was correlated with the type/extent of UC. PPI network analysis revealed common target genes and signaling pathways for both miRNAs. Conclusively, serum miR-486 and miR-25 might serve as new biomarkers of IBD, with serum miR-486 could be employed in risk stratification of IBD subtypes and has the ground for clinical utility in CD diagnosis, whereas miR-25 has potential for UC and CD diagnosis.

Association of integrin-ß2 polymorphism and expression with the risk of rheumatoid arthritis and osteoarthritis in Egyptian patients.

BACKGROUND: The genetic architecture of rheumatoid arthritis (RA) and osteoarthritis (OA) are still unclear. Although RA and OA have quite different causes, they share synovial inflammation, risk factors, and some disease-associated genes, including the integrin subunit ß2 (ITGB2)/CD18 gene involved in extracellular matrix interactions and immune cell signaling. However, the functional role of ITGB2 genetic variants, its circulating expression pattern, and their clinical usefulness in RA and OA remain unexplored. Our study appraised the association of ITGB2 rs2070946 single nucleotide polymorphism with the vulnerability to RA and OA and its influence on ITGB2 mRNA expression, along with the potential of serum ITGB2 expression in RA and OA diagnosis. METHODS: This study included 70 RA patients, 70 primary OA patients, and 60 healthy volunteers. Genotyping and gene expression analysis were performed using qPCR. Bioinformatics analysis was employed to construct the protein-protein interaction (PPI) network of ITGB2. RESULTS: Serum ITGB2 mRNA expression was upregulated in both RA and OA compared to healthy controls. ITGB2 rs2070946 was associated with escalating risk of both diseases. RA patients harboring the rs2070946 CC or TC + CC genotypes had higher serum ITGB2 expression than the TT genotype carriers. Likewise, OA patients having the minor homozygote CC genotype had higher serum ITGB2 expression than those carrying the TT, TC or TT + TC genotypes. Serum ITGB2 expression showed profound diagnostic potential for RA and OA in receiver-operating characteristic analysis. In RA, serum ITGB2 expression positively correlated with rheumatoid factor and disease activity score 28 (DAS28). The ITGB2-PPI network enriched in cell-cell adhesion, ICAM-3 receptor activity, T-cell activation, leukocyte adhesion, complement binding, and NF-κB, tumor necrosis factor, and interleukin signaling pathways. CONCLUSION: These findings embrace the impact of ITGB2 rs2070946 as a novel genetic biomarker of both RA and OA, which could alter the ITGB2 expression. Serum ITGB2 expression could aid in timely diagnosis of RA and OA.

Screening and evaluation of TBX20 and CITED2 mutations in children with congenital cardiac septal defects: Correlation with cardiac troponin T and caspase-3.

Congenital cardiac septal defect (CCSD) is the main type of congenital heart disease and owns a very high mortality rate among newborns. CCSD is controlled by specific transcription factors, including T-box transcription factor 20 (TBX20) and Cbp/P300 interacting transactivator with Glu/Asp rich carboxy-terminal domain 2 (CITED2) which are key molecular actors in heart development. Here, we screened for mutations in TBX20 and CITED2 genes in Egyptian children with CCSD and assessed their association with CCSD susceptibility and with cardiac troponin T (cTnT) and the apoptotic marker caspase-3 as biochemical markers for CCSD. Thirty unrelated newborns and children affected with CCSD and 30 matched healthy controls with no personal history of cardiac diseases were recruited. Selection criteria were children (<18 years) with any age diagnosed with CCSD using ECHO. Mutational analysis and genotyping were done using PCR-Sanger DNA sequencing technique. Serum cTnT and caspase-3 were analyzed using ELISA. Sequencing analysis identified 2 TBX20 variants (c.766T>C and c.39T>C) in the CCSD and control groups and 2 CITED2 variants (c.12T>C and c.9C>T) in one CCSD patient, while were absent in controls. In silico analysis identified TBX20 c.766T>C (rs3999941) as a missense (F256L) pathogenic variant and the other three variants as synonymous and benign. Compared with controls, TBX20 c.766T>C TC genotype and minor C allele were candidate high-risk factors for CCSD. Besides, serum cTnT and caspase-3 were dramatically elevated in CCSD children compared to controls. TBX20 c.766T>C TC genotype was associated with high cTnT in CCSD children. Conclusively, we advocate TBX20 c.766T>C variant as a potential genetic marker for CCSD which might associate with high cTnT levels. CITED2 genetic variants might have rare incidence among Egyptian CCSD children. Serum cTnT and caspase-3 are useful markers for ascertaining CCSD in children. These data could be exploited in prenatal genetic counseling, pre-implantation genotyping, and therapy of CCSD.

Mitochondrial remodeling in colorectal cancer initiation, progression, metastasis, and therapy: A review.

Colorectal cancer (CRC) is a major health concern with multifactorial pathophysiology representing intense therapeutic challenges. It is well known that deregulation of spatiotemporally-controlled signaling pathways and their metabolic reprogramming effects play a pivotal role in the development and progression of CRC. As such, the mitochondrial role in CRC initiation gained a lot of attention recently, as it is considered the powerhouse that regulates the bioenergetics in CRC. In addition, the crosstalk between microRNAs (miRNAs) and mitochondrial dysfunction has become a newfangled passion for deciphering CRC molecular mechanisms. This review sheds light on the relationship between different signaling pathways involved in metabolic reprogramming and their therapeutic targets, alterations in mitochondrial DNA content, mitochondrial biogenesis, and mitophagy, and the role of polymorphisms in mitochondrial genes as well as miRNAs regulating mitochondrial proteins in CRC initiation, progression, metastasis, and resistance to various therapies.

Dexmedetomidine Attenuates Methotrexate-Induced Neurotoxicity and Memory Deficits in Rats through Improving Hippocampal Neurogenesis: The Role of miR-15a/ROCK-1/ERK1/2/CREB/BDNF Pathway Modulation.

Methotrexate (MTX) is a widely used neurotoxic drug with broad antineoplastic and immunosuppressant spectra. However, the exact molecular mechanisms by which MTX inhibits hippocampal neurogenesis are yet unclear. Dexmedetomidine (Dex), an α2-adrenergic receptor agonist, has recently shown neuroprotective effects; however, its full mechanism is unexplored. This study investigated the potential of Dex to mitigate MTX-induced neurotoxicity and memory impairment in rats and the possible role of the miR-15a/ROCK-1/ERK1/2/CREB/BDNF pathway. Notably, no former studies have linked this pathway to MTX-induced neurotoxicity. Male Sprague Dawley rats were placed into four groups. Group 1 received saline i.p. daily and i.v. on days 8 and 15. Group 2 received Dex at 10 µg/kg/day i.p. for 30 days. Group 3 received MTX at 75 mg/kg i.v. on days 8 and 15, followed by four i.p. doses of leucovorin at 6 mg/kg after 18 h and 3 mg/kg after 26, 42, and 50 h. Group 4 received MTX and leucovorin as in group 3 and Dex daily dosages as in group 2. Bioinformatic analysis identified the association of miR-15a with ROCK-1/ERK1/2/CREB/BDNF and neurogenesis. MTX lowered hippocampal doublecortin and Ki-67, two markers of neurogenesis. This was associated with the downregulation of miR-15a, upregulation of its target ROCK-1, and reduction in the downstream ERK1/2/CREB/BDNF pathway, along with disturbed hippocampal redox state. Novel object recognition and Morris water maze tests demonstrated the MTX-induced memory deficiencies. Dex co-treatment reversed the MTX-induced behavioral, biochemical, and histological alterations in the rats. These neuroprotective actions could be partly mediated through modulating the miR-15a/ROCK-1/ERK1/2/CREB/BDNF pathway, which enhances hippocampal neurogenesis.

Association of LncRNA-PAX8-AS1 and LAIR-2 polymorphisms along with their expression with clinical and subclinical hypothyroidism.

The genetic and epigenetic architecture of clinical and subclinical hypothyroidism remains unclear. We investigated the impact of long noncoding RNA (LncRNA)-PAX8-AS1 and LAIR-2 genetic variants on the susceptibility to clinical and subclinical hypothyroidism, their influence on LncRNA-PAX8-AS1 and LAIR-2 expression and their potential as hypothyroid biomarkers. Hundred clinical hypothyroid patients, 110 subclinical hypothyroid patients, and 95 healthy controls were enrolled. Gene expression analysis and genotyping were performed by qPCR. LAIR-2 protein, a proinflammatory mediator, was tested by ELISA. Serum LncRNA-PAX8-AS1 was downregulated, whereas LAIR-2 mRNA and protein levels were upregulated in clinical and subclinical hypothyroid patients compared to healthy controls. LncRNA-PAX8-AS1 rs4848320 and rs1110839 were associated with increased risk of clinical hypothyroidism. Interestingly, both SNPs were associated with differential expression of serum LncRNA-PAX8-AS1 among clinical hypothyroid patients. LAIR-2 rs2287828 was associated with elevated risk of both clinical and subclinical hypothyroidism. Harboring the rs2287828 T allele augmented the LAIR-2 mRNA expression among clinical hypothyroid patients, while elevated both LAIR-2 mRNA and protein levels in subclinical hypothyroid patients. The rs4848320-rs1110839-rs2287828 TTT, CTT, and CGT haplotypes were associated with increased hypothyroid risk. Surprisingly, serum LncRNA-PAX8-AS1 and LAIR-2 mRNA expression demonstrated superior diagnostic accuracy for clinical hypothyroidism and turned out as independent predictors in the multivariate analysis. Conclusively, LncRNA-PAX8-AS1 and LAIR-2 genetic variants are novel genetic biomarkers of hypothyroidism that could alter the LncRNA-PAX8-AS1 and LAIR-2 expression. LncRNA-PAX8-AS1 and LAIR-2 expression profiles have the potential as effective diagnostic and prognostic indicators of hypothyroidism.

Repaglinide Elicits a Neuroprotective Effect in Rotenone-Induced Parkinson's Disease in Rats: Emphasis on Targeting the DREAM-ER Stress BiP/ATF6/CHOP Trajectory and Activation of Mitophagy.

Repaglinide, a meglitinide insulinotropic antidiabetic, was unraveled as a promising therapeutic agent for Huntington's disease by targeting the neuronal calcium sensor downstream regulatory element antagonist modulator (DREAM). However, its mechanistic profile in Parkinson's disease (PD) especially its impact on endoplasmic reticulum (ER) stress, mitophagy, and their interconnections is poorly elucidated. This study is the first to examine the neuroprotective potential of repaglinide in rotenone-induced PD in rats by exploring its effects on DREAM, BiP/ATF6/CHOP ER stress pathway, apoptosis, mitophagy/autophagy, oxidative stress, astrogliosis/microgliosis, and neuroinflammation. Male Wistar rats were randomly assigned to four groups: groups 1 and 2 received the vehicle or repaglinide (0.5 mg/kg/day p.o). Groups 3 and 4 received rotenone (1.5 mg/kg/48 h s.c) for 21 days; meanwhile, group 4 additionally received repaglinide (0.5 mg/kg/day p.o) for 15 days starting from day 11. Interestingly, repaglinide lessened striatal ER stress and apoptosis as evidenced by reduced BiP/ATF6/CHOP and caspase-3 levels; however, it augmented striatal DREAM mRNA expression. Repaglinide triggered the expression of the mitophagy marker PINK1 and the autophagy protein beclin1 and alleviated striatal oxidative stress through escalating catalase activity. In addition, repaglinide halted astrocyte/microglial activation and neuroinflammation in the striatum as expressed by reducing glial fibrillary acidic protein (GFAP) and ionized calcium-binding adaptor protein 1 (Iba1) immunostaining and decreasing interleukin (IL)-6 and IL-1ß levels. Repaglinide restored striatum morphological alterations, intact neuron count, and neurobehavioral motor performance in rats examined by an open field, grip strength, and footprint gait analysis. Conclusively, repaglinide modulates the DREAM-ER stress BiP/ATF6/CHOP cascade, increases mitophagy/autophagy, inhibits apoptosis, and lessens oxidative stress, astrocyte/microglial activation, and neuroinflammation in PD.

Cromolyn chitosan nanoparticles reverse the DNA methylation of RASSF1A and p16 genes and mitigate DNMT1 and METTL3 expression in breast cancer cell line and tumor xenograft model in mice.

BACKGROUND: Developing epigenetic drugs for breast cancer (BC) remains a novel therapeutic approach. Cromolyn is a mast cell stabilizer emerging as an anticancer drug; its encapsulation in chitosan nanoparticles (CSNPs) improves its effect and bioavailability. However, its effect on DNA and RNA methylation machineries has not been previously tackled. METHODS: The possible anticancer effect of cromolyn CSNPs and its potential as an epigenetic drug was investigated in vitro using MCF-7 human BC cell line and in vivo using Ehrlich ascites carcinoma-xenograft model in mice symbolizing murine mammary adenocarcinoma. Mice were injected with a single dose of Ehrlich ascites carcinoma cells subcutaneously for the induction of tumor mass, and then randomized into three groups: control, cromolyn CSNPs (equivalent to 5 mg cromolyn/kg, i.p.) and plain CSNPs twice/week for 2 weeks. RESULTS: Cromolyn CSNPs showed prominent anticancer effect in MCF-7 cells by reducing the cell viability percent and enhancing DNA damage in the comet assay demonstrating its apoptotic actions. Mechanistically, cromolyn CSNPs influenced potential epigenetic processes through mitigating DNA methyltransferase 1 (DNMT1) expression, reversing the hypermethylation pattern of the tumor suppressor RASSF1A and p16 genes and attenuating the expression of the RNA N6-methyladenosine writer, methyltransferase-like 3 (METTL3). Cromolyn CSNPs diminished ERK1/2 phosphorylation, a possible arm influencing DNMT1 expression. In vivo, cromolyn CSNPs lessened the tumor volume and halted DNMT1 and METTL3 expression in Ehrlich carcinoma mice. CONCLUSIONS: Cromolyn CSNPs have the premise as an epigenetic drug through inhibiting ERK1/2 phosphorylation/DNMT1/DNA methylation and possibly impacting the RNA methylation machinery via mitigating METTL3 expression.

Empagliflozin alleviates endoplasmic reticulum stress and augments autophagy in rotenone-induced Parkinson's disease in rats: Targeting the GRP78/PERK/eIF2α/CHOP pathway and miR-211-5p.

Empagliflozin, a selective sodium-glucose co-transporter-2 inhibitor, has been demonstrated to provide additional non-glycemic benefits, including neuroprotection. Endoplasmic reticulum (ER) stress is a key player in neurodegeneration and occurs at the crossroads of other pathologic mechanisms; however, its role in the pathogenesis of Parkinson's disease (PD) is still elusive. miR-211-5p regulates neuronal differentiation and viability and was predicted to target CHOP, a downstream effector in the ER stress pathway. For the first time, this study investigated the possible neuroprotective effect of empagliflozin in a rotenone-induced rat model of PD from the perspective of ER stress. Rotenone (1.5 mg/kg) was administered subcutaneously every other day for 3 weeks. Meanwhile, the treated group received empagliflozin 10 mg/kg/day orally for 15 consecutive days post-PD induction. On the molecular level, the ER stress pathway components; GRP78, total and phosphorylated PERK, eIF2α and CHOP, along with miR-211-5p expression were upregulated in the striatum of rotenone-injected rats. Concurrently, the untreated rats showed elevated striatal α-synuclein levels along with diminished autophagy and the proteasome system as evidenced by reduced beclin-1 protein and ELF2/NERF mRNA expression levels. The rotenone-induced striatal oxidative stress and neuroinflammation were expressed by reduced catalase activity and elevated interleukin (IL)-1ß levels. miR-211-5p was positively correlated with PERK/eIF2α/CHOP, IL-1ß and α-synuclein, while negatively correlated with ELF2/NERF, beclin-1 and catalase activity. Empagliflozin treatment showed a restorative effect on all biochemical alterations and improved the motor function of rats tested by open field, grip strength and footprint gait analysis. In the histopathological examination, empagliflozin increased the intact neuron count and attenuated astrogliosis and microgliosis by reducing the glial fibrillary acidic protein and ionized calcium-binding adaptor protein 1 immunostaining. Conclusively, these results emphasize the neurotherapeutic impact of empagliflozin in PD by moderating the GRP78/PERK/eIF2α/CHOP ER stress pathway, downregulating miR-211-5p, resolving oxidative stress, lessening astrocyte/microglial activation and neuroinflammation, along with augmenting autophagy.

Exendin-4 enhances osteogenic differentiation of adipose tissue mesenchymal stem cells through the receptor activator of nuclear factor-kappa B and osteoprotegerin signaling pathway.

The capability of mesenchymal stem cells (MSCs) to repair bone damage and defects has long been investigated. The receptor activator of nuclear factor-kappa B (RANK), its ligand (RANKL) and the decoy receptor osteoprotegerin (OPG) axis is crucial to keep the equilibrium between osteoblastic and osteoclastic activity. Exendin-4 utilization increased bone formation and enhanced bone integrity. This study aimed to investigate the mentioned axis and determine the effect of exendin-4 upon adipose mesenchymal stem cells (Ad-MSCs) osteogenic differentiation. Ad-MSCs were isolated from rat epididymal fat, followed by characterization and then differentiation into osteocytes both in the presence or absence of exendin-4. Osteogenic differentiation was evaluated by alizarin red staining and the expression of osteogenic markers; using reverse transcriptase-quantitative polymerase chain reaction, western blotting and enzyme-linked immunoassay. MSCs derived from rat epididymal fat were isolated and characterized, along with their differentiation into osteocytes. The differentiated cells were alizarin red-stained, showing increased staining intensity upon addition of exendin-4. Moreover, the addition of exendin-4 elevated the messenger RNA expression levels of osteogenic markers; runt-related transcription factor-2 (RUNX-2), osteocalcin, and forkhead box protein O-1 while reducing the expression of the adipogenic marker peroxisome-proliferator-activated receptor-gamma. Exendin-4 addition elevated OPG levels in the supernatant of osteogenic differentiated cells. Moreover, exendin-4 elevated the protein levels of glucagon-like peptide-1 receptor and RUNX-2, while decreasing both RANK and RANKL. In conclusion, osteogenic differentiation of Ad-MSCs is associated with increased osteoblastic rather than osteoclastic activity. The findings of this study suggest that exendin-4 can enhance Ad-MSCs osteogenic differentiation partially through the RANK/RANKL/OPG axis.

Erythropoietin and Bacillus Calmette-Guérin Vaccination Mitigate 3-Nitropropionic Acid-Induced Huntington-like Disease in Rats by Modulating the PI3K/Akt/mTOR/P70S6K Pathway and Enhancing the Autophagy.

Oxidative stress and mitochondrial dysfunction are among the mechanisms expected to explain the pathogenesis of Huntington's disease. Erythropoietin (EPO) and the Bacillus Calmette-Guérin (BCG) vaccine have neuroprotective effects against neurodegenerative diseases; however, the full mechanisms of their action are currently unclear. Here, for the first time, we investigated the neuroprotective effect of BCG vaccination in Huntington-like disease induced by 3-nitropropionic acid (3-NP) and its combination with EPO. Male Wistar rats were randomized into five groups: saline-treated control; 3-NP group (20 mg/kg/day, i.p.) for 7 days; EPO-treated group (5000 IU/kg/day, i.p.) for 14 days after 3-NP administration; live BCG vaccine prophylactic group (5000 cfu/g, i.p.) 10 days prior to 3-NP administration; and live BCG vaccine (5000 cfu/g, i.p.) 10 days before 3-NP administration, followed by EPO treatment (5000 IU/kg/day, i.p.) for 14 days. In a histopathological examination, striatum neurodegeneration was evidenced in the 3-NP injected rats. Administration of 3-NP elevated the levels of p-PI3K, p-Akt, p-mTOR, p-P70S6K, BAX, malondialdehyde, nitric oxide, and cytochrome oxidase while reduced the levels of BCL-2, superoxide dismutase, reduced glutathione, and the autophagy marker microtubule-associated protein light chain 3 in the striatum. EPO and BCG ameliorated the biochemical, histopathological, and behavioral derangements induced by 3-NP, with prominent neuroprotection observed in rats administered the BCG prophylactic combined with EPO treatment. These results highlight the role played by EPO and BCG in the management of 3-NP-induced Huntington-like disease by inhibiting the PI3K/Akt/mTOR/P70S6K pathway and enhancing the autophagy.

Uncovering serum placental-related non-coding RNAs as possible biomarkers of preeclampsia risk, onset and severity revealed MALAT-1, miR-363 and miR-17.

New predictors that could boost early detection of preeclampsia (PE) and prognosticate its severity are urgently needed. We examined serum miR-17, miR-363, MALAT-1 and HOTAIR as potential biomarkers of PE risk, onset and severity. This prospective study included 160 pregnant females; 82 PE cases and 78 healthy pregnancies. Serum samples were collected between 20 to 40 weeks of gestation. Early-onset PE was defined as developing clinical manifestations at ≤ 34 gestational weeks. Severe PE was defined as systolic blood pressure ≥ 160 mmHg and/or diastolic blood pressure ≥ 110 mmHg and proteinuria (≥ 2 g/24 h or ≥ 2+ dipstick). Selection of PE-related non-coding RNAs and functional target gene analysis were conducted using bioinformatics analysis. Expression profiles were assessed by RT-qPCR. Serum miR-363 and MALAT-1 were downregulated, meanwhile miR-17 was upregulated, and HOTAIR was not significantly altered in PE compared with healthy pregnancies. miR-17 was elevated while miR-363 and MALAT-1 were reduced in severe versus mild PE. miR-363 was lower in early-onset versus late-onset PE. MALAT-1, miR-17 and miR-363 showed diagnostic potential and discriminated severe PE, whereas miR-363 distinguished early-onset PE in the receiver-operating-characteristic analysis. miR-363 and MALAT-1 were significantly associated with early and severe PE, respectively in multivariate logistic analysis. In PE, miR-17 and MALAT-1 were significantly correlated with gestational age (r = - 0.328 and r = 0.322, respectively) and albuminuria (r = 0.312, and r = - 0.35, respectively). We constructed the MALAT-1, miR-363, and miR-17-related protein-protein interaction networks linked to PE. Serum miR-17, miR-363 and MALAT-1 could have utility as new biomarkers of PE diagnosis. miR-363 may be associated with early-onset PE and MALAT-1 downregulation correlates with PE severity.

Agomelatine attenuates alcohol craving and withdrawal symptoms by modulating the Notch1 signaling pathway in rats.

AIM: Alcohol abuse is a significant causative factor of death worldwide. The Notch1 signaling pathway is involved in alcohol tolerance, withdrawal and dependence. Agomelatine is a known antidepressant acting as a melatonin receptor (MT1/2) agonist and a 5-hydroxytryptamine receptor-2C antagonist. However, its effects on alcohol cravings and alcohol withdrawal symptoms have not been investigated. In this study, we assessed the possibility of using agomelatine for the treatment of these symptoms in a rat model of alcoholism and the possible role of Notch1 signaling. MAIN METHODS: We induced alcoholism in rats using a free-choice drinking model for 60 days. From day 61, free-choice was continued until day 82 for the craving model, whereas only water was offered in the withdrawal model. Meanwhile, the treated groups for both models received agomelatine (50 mg/kg/day) orally from day 61 to 82, followed by behavioral, histopathological and biochemical assessment. KEY FINDINGS: Agomelatine treatment caused significant decrease in alcohol consumption with a positive effect on anxiety-like behavior in the open field, memory in the Morris water maze and immobility in the forced swim test. Moreover, agomelatine induced the expression of Notch1 pathway markers, including Notch1, NICD, CREB, CCNE-2, Hes-1, both total and phosphorylated ERK1/2, MMP9, Per2and RGS-2 in the hippocampal formation. By contrast, NMDAR expression was reduced. Furthermore, agomelatine normalized the serum levels of BDNF, cortisol, dopamine and glutamate which were disrupted by alcohol consumption. SIGNIFICANCE: Based on these findings, agomelatine reversed alcohol cravings and withdrawal symptoms associated with alcohol dependence by modulating the Notch1 signaling pathway.

Association of MALAT1 and PVT1 Variants, Expression Profiles and Target miRNA-101 and miRNA-186 with Colorectal Cancer: Correlation with Epithelial-Mesenchymal Transition.

The influence of PVT1 and MALAT1 variants on colorectal cancer (CRC) susceptibility and their impact on PVT1/miRNA-186/epithelial-mesenchymal transition (EMT) and MALAT1/miRNA-101/EMT axes in CRC are unknown. We investigated the influence of PVT1 rs13255292 and MALAT1 rs3200401 on the risk of CRC and adenomatous polyps (AP), their impact on the long noncoding RNAs PVT1 and MALAT1 expression and their target miRNA-186, miRNA-101/E-cadherin pathways, along with their potential as early CRC biomarkers. Overall, 280 individuals were recruited: 140 patients with CRC, 40 patients with AP, and 100 healthy volunteers. Genotyping and serum expression profiles were assessed using qPCR. The EMT biomarker, E-cadherin, was measured by ELISA. rs3200401 was associated with increased CRC risk, whereas rs13255292 was protective. Serum PVT1 and MALAT1 were upregulated in CRC and AP patients versus healthy controls, whereas, miRNA-186, miRNA-101 and E-cadherin were downregulated in CRC versus non-CRC groups. MALAT1 showed superior diagnostic potential for CRC and predicted CRC risk among non-CRC groups in the multivariate logistic analysis. PVT1, MALAT1, miRNA-186 and miRNA-101 levels were correlated with E-cadherin, tumor stage, lymph node and distant metastasis. E-cadherin was lost in metastatic vs. non-metastatic CRC. rs3200401CC genotype carriers showed higher E-cadherin levels than CC + CT carriers. rs3200401 was correlated with lymph node status. For the first time, rs13255292 and rs3200401 are potential genetic CRC predisposition markers, with rs3200401 possibly impacting the EMT process. Serum PVT1, MALAT1, miRNA-186 and miRNA-101 are novel non-invasive diagnostic biomarkers that could improve the clinical outcome of CRC.

Linagliptin mitigates experimental inflammatory bowel disease in rats by targeting inflammatory and redox signaling.

AIMS: Dipeptidyl peptidase-4 (DPP-4) has been involved in the pathogenesis of inflammatory bowel diseases (IBD), yet the underlying mechanisms remain inconclusive. The present study aimed to investigate the potential of linagliptin, a potent/selective DPP-4 inhibitor with marked anti-inflammatory actions, to attenuate trinitrobenzene sulfonic acid (TNBS)-evoked colitis in rats; an experimental model of IBD, and the implicated molecular mechanisms. This may add to the clinical utility of linagliptin for the management of patients with coexisting IBD and diabetes mellitus. Notably, no former studies have linked JAK2/STAT3, HMGB1/NF-κB, and Nrf2/HO-1 signaling in TNBS-evoked colitis. MATERIALS AND METHODS: Western blotting and ELISA were used to determine the levels of target signals. KEY FINDINGS: Administration of linagliptin (1.5 mg/kg; p.o.) mitigated the colitis severity via diminishing the disease activity index, colon weight/length ratio, and macroscopic scores. Linagliptin also lowered the colonic histologic scores and leukocyte invasion. Notably, linagliptin inhibited the colonic DPP-4 activity and upregulated the expression of intestinotrophic GLP-2 without incurring hypoglycemia in animals. Linagliptin curbed inflammation through the suppression of colonic IL-6, TNF-α, and myeloperoxidase and upregulation of IL-10. It also inhibited the IL-6/JAK2/STAT3 pathway via downregulating p-JAK2/JAK2 and p-STAT3/STAT3 protein expression and HMGB1/RAGE/NF-κB cascade through lowering HMGB1, RAGE, and p-NF-κB p65/NF-κB p65 protein expression. In the context of mucosal oxidative stress, linagliptin diminished lipid peroxides and augmented GSH, GPx, and total antioxidant capacity. It also activated Nrf2/HO-1 pathway via upregulating Nrf2 and HO-1 protein expression. SIGNIFICANCE: Linagliptin shows a promise for the management of IBD via targeting IL-6/JAK2/STAT3, HMGB1/RAGE/NF-κB, and Nrf2/HO-1 pathways.