M1 Expression Research Articles

Milk Formula Enriched with Sodium Butyrate Influences Small Intestine Contractility in Neonatal Pigs.

Butyrate, a by-product of gut bacteria fermentation as well as the digestion of fat in mother’s milk, exerts a wide spectrum of beneficial effects in the gastrointestinal tissues. The present study aimed to determine the effects of sodium butyrate on small intestine contractility in neonatal piglets. Piglets were fed milk formula alone (group C) or milk formula supplemented with sodium butyrate (group B). After a 7-day treatment period, isometric recordings of whole-thickness segments of the duodenum and middle jejunum were obtained by electric field stimulation under the influence of increasing doses of Ach (acetylocholine) in the presence of TTX (tetrodotoxin) and atropine. Moreover, structural properties of the intestinal wall were assessed, together with the expression of cholinergic and muscarinic receptors (M1 and M2). In both intestinal segments (duodenum and middle jejunum), EFS (electric field stimulation) impulses resulted in increased contractility and amplitude of contractions in group B compared to group C. Additionally, exposure to dietary butyrate led to a significant increase in tunica muscularis thickness in the duodenum, while mitotic and apoptotic indices were increased in the middle jejunum. The expression of M1 and M2 receptors in the middle jejunum was significantly higher after butyrate treatment. The results indicate increased cholinergic signaling and small intestinal growth and renewal in response to feeding with milk formula enriched with sodium butyrate in neonatal piglets.

The Construction of ITP Diagnostic Modeling Based on the Expressions of Hub Genes Associated with M1 Polarization of Macrophages.

PurposePrimary immune thrombocytopenia (ITP) is an immune disease with a diagnosis of exclusion, since no validated biomarkers have been identified. In this study, we explored biomarkers associated with the development of ITP from an immune perspective to inform the clinical diagnosis.Patients and MethodsDifferentially expressed genes (DEGs) between normal and ITP samples were analyzed using limma package. Random forest algorithm and LASSO regression were further used to screen for DEGs associated with ITP. The expression of these hub genes was validated by PCR. The relationship between DEGs and immunity was explored by enrichment analysis. Immune cell infiltration in ITP was analyzed by CIBERSORT and ssGSEA, and the relationship between DEGs and infiltrating immune cells was analyzed by Spearman’s rank correlation analysis. Finally, a diagnostic model related to DEGs was constructed by the neural network, and its efficiency was detected by the ROC curve.ResultsAfter screening the GEO database and validation by PCR analysis, The expression of CTH and TAF8 were higher and while OSBP2 expression was lower in ITP patients compared to normal subjects (P<0.05). GO enrichment analysis showed that these DEGs were associated with inflammatory immune-related diseases, and KEGG analysis showed that they mainly regulated signaling pathways such as JAK-STAT. CIBERSORT and ssGSEA analyses showed that these DEGs were mainly associated with macrophage M1 polarization. The expression of CTH and TAF8 were positively correlated with M1 expression, while OSBP2 was negatively correlated with M1 expression. The ROC curve showed high accuracy of the neural network model [AUC= 0.939, 95% CI (0.8–1)].ConclusionOur results suggest that CTH, TAF8, and OSBP2 can be used as effective diagnostic biomarkers of ITP.

Minocycline attenuates cholinergic dysfunction and neuro-inflammation-mediatedcognitive impairmentin scopolamine-induced Alzheimer's rat model.

Minocycline, a semisynthetictetracycline-derivedantibiotic, has various pharmacological effect such as anti-inflammatory, anti-oxidative stress, and anti-apoptotic effects. The current study investigated the involvement of neuro-inflammatory, oxidative stress, and cholinergic markers in neuroprotection by minocycline against scopolamine-induced brain damage. Minocycline was administered (oral, 10, 15, and 30mg/kg, daily) to groups of amnesic rats for 21days. Passive avoidance memory and spatial learning and memory were assessed. Following that, oxidative stress, cholinergic function, and neuro-inflammation markers were evaluated in the brain tissue. According to our biochemical data, treatment of the scopolamine-injured rats with minocycline decreased the levels of malondialdehyde and acetylcholinesterase (AChE) as well as mRNA expression of AChE and neuro-inflammation markers (tumor necrosis factor-α, interleukin (IL)-1β, IL-6). It also increased the total thiol levels and superoxide dismutase activity as well as mRNA expression of cholinergic receptor M1 (ChRM1). Moreover, minocycline modified distance and latencies in Morris water maze, prolonged latency to enter the black zone and light time while decreasing time spent and frequency of entries to darkness. Taken together, the data indicate that treatment with minocycline improved memory dysfunction mediated possibly through restoring AChE and ChRM1 levels, oxidant/antioxidant balance, as well as inhibiting inflammatory responses.

Muscarinic receptor regulation of chronic pain-induced atrial fibrillation.

Atrial fibrillation (AF), one of the most common arrhythmias, is associated with chronic emotional disorder. Chronic pain represents a psychological instability condition related to cardiovascular diseases, but the mechanistic linkage connecting chronic pain to AF occurrence remains unknown. Wild-type C57BL/6J male mice were randomly divided into sham and chronic pain groups. Autonomic nerve remodeling was reflected by the increased atrial parasympathetic tension and muscarinic acetylcholine receptor M2 expression. AF susceptibility was assessed through transesophageal burst stimulation in combination with electrocardiogram recording and investigating AERP in Langendorff perfused hearts. Our results demonstrated the elevated protein expression of muscarinic acetylcholine receptor M2 in the atria of mice subjected to chronic pain stress. Moreover, chronic pain induced the increase of atrial PR interval, and atrial effective refractory periods as compared to the sham group, underlying the enhanced susceptibility of AF. Thus, autonomic cholinergic nerve may mediate mice AF in the setting of chronic pain.

Knockdown MTDH Inhibits Glioma Proliferation and Migration and Promotes Apoptosis by Downregulating MYBL2.

Glioma is a malignant tumor that often occurs in the adult central nervous system. Metadherin/astrocyte-elevated gene-1 (MTDH) is involved in the development of cancer, but its relationship with glioma remains unclear. This study is aimed at clarifying the role of MTDH in glioma. GEPIA was employed to find the difference of the expression level of MTDH and MYB protooncogene-like 2 (MYBL2) in glioma tissues and normal tissues, and real-time quantitative reverse transcription PCR (qRT-PCR) and western blot (WB) were applied to verify the differential gene expression of MTDH and MYBL2 cells. After knocking down of MTDH, the expressions of forkhead box M1 (FoxM1), MTDH, and MYBL2 were detected by WB cells. Cell counting kit 8 (CCK-8) was used to detect cell proliferation, and flow cytometry was applied to measure cell apoptosis. The transwell assay was utilized to investigate the ability of cell migration and invasion. The results showed that MTDH and MYBL2 were overexpressed in glioma cells compared with normal cells. The knockdown of MTDH would inhibit the expression of MYBL2 through decreasing the expression of FoxM1 and further reduce glioma cell proliferation and cell migration and invasion. The present study showed that knockdown of MTDH inhibits glioma proliferation and migration and promotes apoptosis by downregulating MYBL2, which suggests that MTDH is a potential gene in clinical treatment of glioma.

Role of Estrogen Receptor-Positive/Negative Ratios in Regulating Breast Cancer.

The alpha estrogen receptor (ERα) contributes to breast cancer progression and recent guidelines define ER positivity as ≥1% stained cells, and a few tumor tissues show no ERα expression at all or are at 100%. Although ER and aromatase inhibitors are widely used to treat hormone receptor-positive (HR+) breast cancer, their effect on tumor activity at different ERα levels remains unclear. Therefore, we investigated the role of ERα+/ERα− ratios in determining the ERα level. We used ERα stably transfected and wild-type MDA-MB-231 cells (MDA-MB-231Trans−ER and MDA-MB-231WT, respectively) as represented ER+ and ER− cells, respectively, and MCF-7 cells were the positive control. MDA-MB-231Trans−ER and MDA-MB-231WT cells were mixed and cocultured at a ratio of 0%, 20%, 40%, 70%, and 100%. Migration and invasion functions at different cell ratios were evaluated in vitro using the Transwell and scratch test. In a xenograft mouse model, the polarization of the tumor-associated (M2) macrophage and the expression of breast cancer gene 1 (BRCA1), human epidermal growth factor receptor 2 (HER2), vascular endothelial growth factor (VEGF), and tumor necrosis factor (TNF)-α were measured. The results showed that the cell invasion and migration were significantly higher at 40% and 70% than they were at other ratios. Additionally, in vivo, the 70% ERα+/ERα-ratio was a critical indicator of cell activity and cytokine expression. The highest M2 level and expression of VEGR, TNF-α, BRCA1, and HER2 were shown at a ratio of 70%. Moreover, the effects of ERα were not linear in breast cancer, indicating that the ERα status requires continuous monitoring during long-term endocrine treatment. These results indicate that during HR+ breast cancer treatment, the ERα+/ERα− ratio may be a useful predictor and should be evaluated further.

Microglial Dynamics Modulate Vestibular Compensation in a Rodent Model of Vestibulopathy and Condition the Expression of Plasticity Mechanisms in the Deafferented Vestibular Nuclei.

Unilateral vestibular loss (UVL) induces a vestibular syndrome composed of posturo-locomotor, oculomotor, vegetative, and perceptivo-cognitive symptoms. With time, these functional deficits progressively disappear due to a phenomenon called vestibular compensation, known to be supported by the expression in the deafferented vestibular nuclei (VNs) of various adaptative plasticity mechanisms. UVL is known to induce a neuroinflammatory response within the VNs, thought to be caused by the structural alteration of primary vestibular afferents. The acute inflammatory response, expressed in the deafferented VNs was recently proven to be crucial for the expression of the endogenous plasticity supporting functional recovery. Neuroinflammation is supported by reactive microglial cells, known to have various phenotypes with adverse effects on brain tissue. Here, we used markers of pro-inflammatory and anti-inflammatory phenotypes of reactive microglia to study microglial dynamics following a unilateral vestibular neurectomy (UVN) in the adult rat. In addition, to highlight the role of acute inflammation in vestibular compensation and its underlying mechanisms, we enhanced the inflammatory state of the deafferented VNs using systemic injections of lipopolysaccharide (LPS) during the acute phase after a UVN. We observed that the UVN induced the expression of both M1 proinflammatory and M2 anti-inflammatory microglial phenotypes in the deafferented VNs. The acute LPS treatment exacerbated the inflammatory reaction and increased the M1 phenotype while decreasing M2 expression. These effects were associated with impaired postlesional plasticity in the deafferented VNs and exacerbated functional deficits. These results highlight the importance of a homeostatic inflammatory level in the expression of the adaptative plasticity mechanisms underlying vestibular compensation. Understanding the rules that govern neuroinflammation would provide therapeutic leads in neuropathologies associated with these processes.

Qingchang Wenzhong Decoction Alleviates DSS-Induced Inflammatory Bowel Disease by Inhibiting M1 Macrophage Polarization In Vitro and In Vivo.

An imbalance of macrophage M1/M2 polarization significantly influences the pathogenesis of inflammatory bowel disease. Qingchang Wenzhong decoction (QCWZD) has a proven therapeutic effect on patients with inflammatory bowel disease (IBD) and can significantly inhibit the inflammatory response in mice with colitis. However, its effect on macrophages during IBD treatment remains nebulous. Aim of the Study. Explore the mechanism underlying QCWZD effects in a dextran sulfate sodium (DSS)-induced colitis mouse model in vivo and RAW264.7 cell in vitro by observing macrophage polarization dynamics. The main active components of QCWZD were determined using high-performance liquid chromatography. Surface marker expression on M1-type macrophages was analyzed using flow cytometry and immunofluorescence. The effect on inducible nitric oxide synthase (iNOS), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) released by M1 type macrophages was determined using ELSA and RT-PCR. The expression of key proteins in the JAK2/STAT3 signaling pathway was analyzed using western blotting. QCWZD cytotoxicity in macrophages was measured using CCK8 and Annexin V-FITC/PI assays. The main active components of QCWZD were berberine chloride, coptisine chloride, epiberberine chloride, gallic acid, ginsenoside Rg1, ginsenoside Rb1, indigo, indirubin, notoginsenoside R1, palmatine chloride, and 6-curcumin. QCWZD markedly alleviated DSS-induced colitis in mice, as revealed by the rescued weight loss and disease activity index, attenuated the colonic shortening and mucosal injury associated with the inhibition of M1 macrophage polarization and expression of related cytokines, such as IL-6 and TNF-α, in vivo and in vitro. Furthermore, QCWZD decreased the iNOS, JAK2, and STAT3 levels in vivo and in vitro, regulating the JAK2/STAT3 signaling pathway. QCWZD administration improves intestinal inflammation by inhibiting M1 macrophage polarization. The JAK2/STAT3 signaling pathway may mediate the effects of QCWZD on M1 macrophage polarization in colitis treatment. This study presents a novel macrophage-mediated therapeutic strategy for the treatment of IBD.

PD 102807 Induces M3 mAChR-Dependent GRK-/Arrestin-Biased Signaling in Airway Smooth Muscle Cells.

G protein-coupled receptors (GPCRs) not only are turned on or off to control canonical G protein signaling but also may be fine-tuned to promote qualitative/biased signaling. Qualitative signaling by M3 muscarinic acetylcholine receptors (mAChRs) has been proposed, but its impact on physiologic systems remains unclear, and currently no biased M3 mAChR ligands have been described. Herein, we identify PD 102807 as a biased M3 ligand and delineate its signaling and function in human airway smooth muscle (ASM) cells. PD 102807 induced M3-mediated β-arrestin recruitment but not calcium mobilization. PD 102807 inhibited methacholine (MCh)-induced calcium mobilization in (M3-expressing) ASM cells. PD 102807 induced phosphorylation of AMP-activated protein kinase (AMPK) and the downstream effector acetyl-coenzyme A carboxylase (ACC). PD 102807- induced phosphorylated (p)-AMPK levels were greatly reduced in ASM cells with minimal M3 expression and were not inhibited by the Gq inhibitor YM-254890. Induction of p-AMPK and p-ACC was inhibited by β-arrestin 1 or GRK2/3 knockdown. Similarly, MCh induced phosphorylation of AMPK/ACC, but these effects were Gq dependent and unaffected by GRK2/3 knockdown. Consistent with the known ability of AMPK to inhibit transforming growth factor β (TGF-β)-mediated functions, PD 102807 inhibited TGF-β-induced SMAD-Luc activity, sm-α-actin expression, actin stress fiber formation, and ASM cell hypercontractility. These findings reveal that PD 102807 is a biased M3 ligand that inhibits M3-transduced Gq signaling but promotes Gq protein-independent, GRK-/arrestin-dependent, M3-mediated AMPK signaling, which in turn regulates ASM phenotype and contractile function. Consequently, biased M3 ligands hold significant promise as therapeutic agents capable of exploiting the pleiotropic nature of M3 signaling.

Local and systemic inflammation triggers different outcomes of tumor growth related to infiltration of anti-tumor or pro-tumor macrophages.

Background:Previous evidence suggests inflammation may be a double-edged sword with cancer-promoting and cancer suppressing function. In this study, we explore the impact of local and systemic inflammation on cancer growth.Methods:Female BALB/C mice were subcutaneously implanted with foreign body (plastic plates) to build up a local inflammation and intraperitoneally injected with PolyIC or lipopolysaccharides (LPS) to build up a systemic inflammation, followed by subcutaneous injection of 5 × 105 colon cancer cells. Immunohistochemistry and enzyme linked immunosorbent assay were utilized to detect the Ki67 and interleukin (IL) 6, IL-1β, and monocyte chemoattractant protein-1 expression in the tumor tissues and serum, respectively. The distributions of immune cells and expression of toll-like receptors (TLRs) were evaluated by flow cytometry (FCM) and quantitative real time-polymerase chain reaction.Results:The results showed that local inflammation induced by foreign body implantation suppressed tumor growth with decreased tumor weight (P = 0.001), volume (P = 0.004) and Ki67 index (P < 0.001). Compared with the control group, myeloid-derived suppressive cells sharply decreased (P = 0.040), while CD4+ T cells slightly increased in the tumor tissues of the group of foreign body-induced local inflammation (P = 0.035). Moreover, the number of M1 macrophages (P = 0.040) and expression of TLRs, especially TLR3 (P < 0.001) and TLR4 (P < 0.001), were significantly up-regulated in the foreign body group. Contrarily, tumor growth was significantly promoted in LPS or PolyIC-induced systemic inflammation (P = 0.009 and 0.006). FCM results showed M1 type macrophages (P = 0.017 and 0.006) and CD8+ T cells (P = 0.031 and 0.023) were decreased, while M2 type macrophages (P = 0.002 and 0.007) were significantly increased in tumor microenvironment of LPS or PolyIC-induced systemic inflammation group. In addition, the decreased expression of TLRs was detected in LPS or PolyIC group.Conclusions:The foreign body-induced local inflammation inhibited tumor growth, while LPS or PolyIC- induced systemic inflammation promoted tumor growth. The results suggested that the different outcomes of tumor growth might be attributed to the infiltration of anti-tumor or pro-tumor immune cells, especially M1 or M2 type macrophages into tumor microenvironment.

Sulforaphane diminishes moonlighting of pyruvate kinase M2 and interleukin 1β expression in M1 (LPS) macrophages.

Murine macrophages activated by the Toll-like receptor 4 agonist lipopolysaccharide (LPS) polarize to the M1 type by inducing proinflammatory marker proteins and changing their energy metabolism to increased aerobic glycolysis and reduced respiration. We here show that the aliphatic isothiocyanate sulforaphane (Sfn) diminishes M1 marker expression (IL-1β, IL-6, TNF-α, iNOS, NO, and ROS) and leads to highly energetic cells characterized by both high glycolytic and high respiratory activity as assessed by extracellular flux analysis. Focusing on a potential connection between high glycolytic activity and low IL-1β expression in M1 (LPS/Sfn) macrophages, we reveal that Sfn impedes the moonlighting function of pyruvate kinase M2 (PKM2) in M1 macrophages. Sfn limits mono/dimerization and nuclear residence of PKM2 accompanied by reduced HIF-1α levels, Stat3 phosphorylation at tyrosine 705, and IL-1β expression while preserving high levels of cytosolic PKM2 tetramer with high glycolytic enzyme activity. Sfn prevents glutathionylation of PKM2 in LPS-stimulated macrophages which may account for the reduced loss of PKM2 tetramer. Overall, we uncover PKM2 as a novel affected hub within the anti-inflammatory activity profile of Sfn.

The distribution pattern of M2 and Adrenergicα2 receptors on inferior colliculi in male newborns of diabetic rats

AimsDespite the high prevalence of diabetes in the world, its possible effects throughut pregnancy on neonatal auditory nervous system development are still unknown. In the present research, maternal diabetes’ impact on the M2 and Adrenergicα2 receptors expression in the inferior colliculus (IC) of male newborn rats was investigated. Main MethodsFemale rats were grouped into three: sham, insulin-treated diabetic, and diabetic. Diabetes was induced through streptozotocin (STZ) injection as one dose intraperitoneally (65 mg/kg). After mating and delivery, male rats were euthanized on P0, P7, and P14. Immunohistochemistry (IHC) was used to study the distribution pattern of receptors. Key FindingsThe present study indicated that the expression of M2 receptors in the diabetic group was significantly increased in pairwise comparisons in the sham and diabetic treated with insulin groups (P < 0.001, each). The highest M2 expression was for the diabetic group on P14 and the lowest one was for the sham group on P0. The Adrenergicα2a receptors expression in the diabetic group was significantly reduced in pairwise comparisons in the sham and diabetic treated with insulin groups (P < 0.001, each). The highest Adrenergicα2a expression was for the sham group on P14 and the lowest one was for the diabetic group on P0. There was no significant difference between the sham and insulin groups regarding all receptors expression. SignificanceThis study demonstrated a time-dependent significant decrease in Adrenergicα2a but a time-dependent significant increase in M2 receptors expression.

Impaired sweating in patients with cholinergic urticaria is linked to low expression of acetylcholine receptor CHRM3 and acetylcholine esterase in sweat glands.

BackgroundCholinergic urticaria (CholU), a frequent form of chronic inducible urticaria, is characterized by itchy wheals and angioedema in response to sweating. As of now, the rate and pathophysiological relevance of impaired sweating in patients with CholU are ill-defined.AimTo assess in CholU patients the rate and extent of impaired sweating and its links to clinical and pathophysiological features of CholU.Patients and methodsWe assessed sweating in patients with CholU (n = 13) subjected to pulse-controlled ergometry (PCE) provocation testing. Pre- and post-PCE biopsies of lesional (L) and non-lesional (NL) skin were analyzed for the expression of acetylcholine receptor M3 (CHRM3) and acetylcholine esterase (ACh-E) by quantitative histomorphometry and compared to those of healthy control subjects (HCs). CholU patients were assessed for disease duration and severity as well as other clinical features.ResultsOf the 13 patients with CholU, 10 showed reduced sweating in response to PCE provocation, and 3 had severely reduced sweating. Reduced sweating was linked to long disease duration and high disease severity. CholU patients with impaired sweating responses showed reduced sweat gland epithelial expression of CHRM3 and ACh-E.ConclusionReduced sweating is common in CholU patients, especially in those with long-standing and severe disease, and it can be severe. Reduced expression of CHRM3 and ACh-E may be the cause or consequence of CholU in patients with impaired sweating, and this should be explored by further studies.

HNRNPK alleviates RNA toxicity by counteracting DNA damage in C9orf72 ALS

A ‘GGGGCC’ repeat expansion in the first intron of the C9orf72 gene is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). The exact mechanism resulting in these neurodegenerative diseases remains elusive, but C9 repeat RNA toxicity has been implicated as a gain-of-function mechanism. Our aim was to use a zebrafish model for C9orf72 RNA toxicity to identify modifiers of the ALS-linked phenotype. We discovered that the RNA-binding protein heterogeneous nuclear ribonucleoprotein K (HNRNPK) reverses the toxicity of both sense and antisense repeat RNA, which is dependent on its subcellular localization and RNA recognition, and not on C9orf72 repeat RNA binding. We observed HNRNPK cytoplasmic mislocalization in C9orf72 ALS patient fibroblasts, induced pluripotent stem cell (iPSC)-derived motor neurons and post-mortem motor cortex and spinal cord, in line with a disrupted HNRNPK function in C9orf72 ALS. In C9orf72 ALS/FTD patient tissue, we discovered an increased nuclear translocation, but reduced expression of ribonucleotide reductase regulatory subunit M2 (RRM2), a downstream target of HNRNPK involved in the DNA damage response. Last but not least, we showed that increasing the expression of HNRNPK or RRM2 was sufficient to mitigate DNA damage in our C9orf72 RNA toxicity zebrafish model. Overall, our study strengthens the relevance of RNA toxicity as a pathogenic mechanism in C9orf72 ALS and demonstrates its link with an aberrant DNA damage response, opening novel therapeutic avenues for C9orf72 ALS/FTD.

Adipose-derived stromal cells reverse insulin resistance through inhibition of M1 expression in a type 2 diabetes mellitus mouse model

BackgroundAdipose tissue inflammation is considered as one of the major mechanisms underlying the pathogenesis of insulin resistance and complications in diabetes. Here, we aimed to study the effects of adipose-derived stromal cells on diabetes-induced insulin resistance and M1 cytokine expression.MethodsStromal vascular fractions (SVFs) purified from the inguinal adipose tissue of diabetic mice were treated with plasma from either nondiabetic (Lepr+/+) or diabetic (Leprdb/db) mice and injected into the inguinal white adipose tissue of Leprdb/db mice.ResultsWe found that diabetic plasma treatment induced, whereas nondiabetic plasma suppressed TNF-α, IL-1β, and dipeptidyl peptidase 4 (DPP4) mRNA expression in SVFs in vitro. Importantly, the injection of nondiabetic plasma-treated SVFs significantly decreased TNF-α, IL-6, IL-1β, CCL2, and IL-33 and induced IL-10 mRNA expression in adipose tissue of Leprdb/db mice in vivo. Furthermore, we observed that nondiabetic plasma-treated SVFs increased mRNA expression of Foxp3 in adipose tissue macrophages and Foxp3 in adipose CD4+ T cells, decreased CD11b+CD11c+ cells in adipose tissue, and suppressed mRNA expression of ICAM-1, FCM3, IL-6, IL-1β, iNOS, TNF-α, and DPP4 as well as protein expression of DPP4 and phosphorylated JNK and NF-κB in the liver of Leprdb/db mice. Moreover, we found that nondiabetic plasma-treated SVFs increased Akt activation following insulin administration and attenuated glucose intolerance in Leprdb/db mice.ConclusionsOur results demonstrate that nondiabetic plasma inhibits M1 but increases M2 cytokine expression in adipose tissue of diabetic mice. Most importantly, our findings reveal that nondiabetic plasma-treated SVFs are capable of mitigating diabetes-induced plasma DPP4 activity, liver inflammation, and insulin resistance and that may be mediated through suppressing M1 cytokines but increasing IL-10 and Tregs in adipose tissue. Altogether, our findings suggest that adipose stromal cell-based therapy could potentially be developed as an efficient therapeutic strategy for the treatment of diabetes.

Bioinformatics Analysis Reveals FOXM1/BUB1B Signaling Pathway as a Key Target of Neosetophomone B in Human Leukemic Cells: A Gene Network-Based Microarray Analysis.

Abnormal expression of Forkhead box protein M1 (FOXM1) and serine/threonine kinase Budding uninhibited by benzimidazoles 1 (BUB1B) contributes to the development and progression of several cancers, including chronic myelogenous leukemia (CML). However, the molecular mechanism of the FOXM1/BUB1B regulatory network and the role of Neosetophomone-B (NSP-B) in leukemia remains unclear. NSP-B, a meroterpenoid fungal secondary metabolite, possesses anticancer potential in human leukemic cells lines; however, the underlying mechanism has not been elucidated. The present study aimed to explore the role of NSP-B on FOXM1/BUB1B signaling and the underlying molecular mechanism of apoptosis induction in leukemic cells. We performed gene expression profiling of NSP-B-treated and untreated leukemic cells to search for differentially expressed genes (DEGs). Interestingly BUB1B was found to be significantly downregulated (logFC -2.60, adjusted p = 0.001) in the treated cell line with the highest connectivity score among cancer genes. Analysis of TCGA data revealed overexpression of BUB1B compared to normal in most cancers and overexpression was associated with poor prognosis. BUB1B also showed a highly significant positive correlation with FOXM1 in all the TCGA cancer types. We used human leukemic cell lines (K562 and U937) as an in vitro study model to validate our findings. We found that NSP-B treatment of leukemic cells suppressed the expression of FOXM1 and BUB1B in a dose-dependent manner. In addition, NSP-B also resulted in the downregulation of FOXM1-regulated genes such as Aurora kinase A, Aurora kinase B, CDK4, and CDK6. Suppression of FOXM1 either by siRNA or NSP-B reduced BUB1B expression and enhanced cell survival inhibition and induction of apoptosis. Interestingly combination treatment of thiostrepton and NSP-B suppressed of cell viability and inducted apoptosis in leukemic cells via enhancing the activation of caspase-3 and caspase-8 compared with single-agent treatment. These results demonstrate the important role of the FOXM1/BUB1B pathway in leukemia and thus a potential therapeutic target.

Protective effects of (R)-enantiomers but not (S)-enantiomers of β2-adrenergic receptor agonists against acute colitis: The role of β2AR

The outcomes of ulcerative colitis (UC) treatment remain unsatisfactory. Salbutamol is a β2-adrenergic receptor (β2AR) agonist that is frequently used to treat human airway diseases, and it is a chiral drug with (RS)-isomers. However, the effects of (RS)-enantiomers of this drug on acute ulcerative colitis remain unknown. The present work determined and compared the effects of different chiral β2AR agonists in acute colitis. Acute colitis was established in mice with 3% dextran sulfate sodium and the mice were orally administered different salbutamol isomers. Body weight loss, colon length, disease activity index (DAI), and colon histopathology were assessed. Inflammatory cytokine levels were detected by ELISA. Colonic biopsies were collected from colitis patients. 16S rDNA amplicon sequencing was carried out to assess the composition and relative abundance of the gut microbiome. The expression of M1 and M2 macrophage markers in the colon were assessed by immunofluorescence staining and Western blotting. The results revealed that (R)-salbutamol prevented body weight loss and colonic shortening, decreased the DAI and histopathological scores, and reduced splenomegaly and inflammatory cytokine levels significantly better than (RS)-salbutamol and (S)-salbutamol. (R)-salbutamol downregulated levels of inflammatory protein in LPS-induced human colon tissue specimens. Furthermore, (R)-salbutamol ameliorated gut dysbiosis and macrophage polarization in mice with colitis. The β2AR antagonist ICI-118551 reversed the effect of (R)-salbutamol in ameliorating acute colitis. Taken together, (R)-salbutamol ameliorated the mice with acute colitis, which can serve as a new candidate or lead compound for UC treatment.

Liver cancer heterogeneity modeled by in situ genome editing of hepatocytes.

Mechanistic study and precision treatment of primary liver cancer (PLC) are hindered by marked heterogeneity, which is challenging to recapitulate in any given liver cancer mouse model. Here, we report the generation of 25 mouse models of PLC by in situ genome editing of hepatocytes recapitulating 25 single or combinations of human cancer driver genes. These mouse tumors represent major histopathological types of human PLCs and could be divided into three human-matched molecular subtypes based on transcriptomic and proteomic profiles. Phenotypical characterization identified subtype- or genotype-specific alterations in immune microenvironment, metabolic reprogramming, cell proliferation, and expression of drug targets. Furthermore, single-cell analysis and expression tracing revealed spatial and temporal dynamics in expression of pyruvate kinase M2 (Pkm2). Tumor-specific knockdown of Pkm2 by multiplexed genome editing reversed the Warburg effect and suppressed tumorigenesis in a genotype-specific manner. Our study provides mouse PLC models with defined genetic drivers and characterized phenotypical heterogeneity suitable for mechanistic investigation and preclinical testing.

Effects of miRNA-149-5p and Platelet-Activating Factor-Receptor Signaling on the Growth and Targeted Therapy Response on Lung Cancer Cells

Accumulating evidence indicates that microRNAs (miRs) play critical roles in essentially all biological processes and their altered expression has been documented in various disease conditions, including human malignancies. Although several cellular mechanisms have been identified in mediating the effects of miRs, the involvement of G-protein-coupled, platelet-activating factor-receptor (PAFR) signaling in miR-149-5p-induced effects on lung cancer growth and therapeutic potential has not been studied. To that end, we first evaluated the functional significance of PAFR and miR-149-5p in A549 and H1299 human non-small cell lung cancer (NSCLC) cell lines. We observed that these tumor lines express endogenous PAFR and miR-149-5p and that PAFR activation by PAF agonist (CPAF) significantly increased, whereas miR-149-5p mimic transfection inhibited cell proliferation in a dose-dependent manner. Interestingly, miR-149-5p mimic significantly attenuated CPAF-mediated increased proliferation of NSCLC cells, as confirmed by miR-149-5p, cyclin D1, and forkhead box protein M1 (FOXM1) expression analysis via qPCR. Our next studies examined PAFR- and miR-149-5p-mediated effects on targeted therapy (i.e., erlotinib and gefitinib) responses. We observed that erlotinib and gefitinib inhibited A549 and H1299 cell survival in a dose- and time-dependent manner, and CPAF significantly blocked this effect. These findings indicate that miR-149-5p blocks PAFR-mediated increased cell proliferation, and PAFR activation attenuates the cytotoxic effects of targeted therapy.

Paternal stress alters synaptic density and expression of GAP-43, GRIN1, M1 and SYP genes in the hippocampus and cortex of offspring of stress-induced male rats

Adverse experiences during pregnancy have a negative impact on the neuronal structure and behavior of offspring, but the effects of a father's life events on the outcome of progeny are scarce. The present study is intended to investigate whether paternal stress affects the offspring brain structure, especially those regions concerned with learning and formation of memory, namely the hippocampus (HC) and prefrontal cortex (PFC), and also the expression of certain genes linked to learning and memory in the offspring. Induced stress to male rats by five stressors, one per day followed by allowing them to mate with the normal, unstressed female. Synaptophysin immunoreactivity was assessed in the tissue sections of the HC and PFC as well as expression of genes concerned with learning and memory was evaluated by RT-PCR in the progeny of stress-received males. The progeny of stressed rats had reduced antisynaptophysin immunoreactivity in the HC and PFC. The synaptic density in HC was less in the A-S (Offspring of male rats who received stress during adulthood) and PA-S (offspring of male rats who received stress during both adolescence and adulthood) than in P-S (offspring of male rats who received stress during adolescence) and C-C (offspring of control) groups. Similar results were observed even in the PFC. The results of post hoc tests proved that the HC and PFC of the progeny of stress-exposed rats exhibited considerably less synaptic density than control (P<0.05), and the levels of expression of GAP-43, GRIN1, M1, and SYP genes in HC and PFC were down-regulated. This study concludes that paternal adverse experiences can affect the offspring's synaptic plasticity and also the genes, which can regulate learning and formation of memory.