Effects of amyloid ß (Aß)42 and Gasdermin D on the progression of Alzheimer's disease in vitro and in vivo through the regulation of astrocyte pyroptosis.

PURPOSE: The study aimed to investigate whether astrocyte pyroptosis, and the subsequent neuroinflammatory response that exerts amyloid ß (Aß) neurotoxic effects, has an effect on endothelial cells, along with the underlying mechanisms. METHODS: In vivo, 5 µL of disease venom was injected into the lateral ventricle of APP/PS1 mice for treatment. Pyroptosis was induced by treating astrocytes with Aß42 in vitro. Small interfering RNA (siRNA) was used to silence caspase-1 and Gasdermin D (GSDMD) mRNA expression. Cell viability was determined using a CCK-8 detection kit. Scanning electron microscopy (SEM), Annexin V/propidium iodide (PI) double staining, RT-qPCR, immunofluorescence, western blotting, and enzyme-linked immunosorbent assay (ELISA) were used to detect cell pyroptosis. The degree of pathological damage to the brain and aortic tissue was assessed by hematoxylin-eosin staining and immunohistochemistry. RESULTS: Aß42 induced astrocyte pyroptosis dependent on the GSDMD/Gasdermin E (GSDME)/Caspase 11/NLRP3 pathway, releasing large amounts of inflammatory factors, such as TNF-α, IL-1α, IL-1ß, and IL-18. Astrocyte pyroptosis caused endothelial cell dysfunction and release of large amounts of vasoconstrictors (ET and vWF). Knockdown of GSDMD reduced astrocyte pyroptosis in the cerebral cortex and hippocampal tissue, decreased the release of inflammatory factors IL-1 ß and IL-18, reduced Aß deposition and tau protein, increased the release of peripheral vasodilator substances (eNOS), and decreased the release of vasoconstrictor substances (ET, vWF), thereby reducing brain tissue damage and vascular injury in APP/PS1 mice. CONCLUSION: Aß42 induced astrocyte pyroptosis, while GSDMD knockout inhibited astrocyte pyroptosis, reduced the release of inflammatory factors, and alleviated brain tissue damage and vascular damage in APP/PS1 mice. Therefore, GSDMD is a novel therapeutic target for Alzheimer's disease. PURPOSE: The study aimed to investigate whether astrocyte pyroptosis, and the subsequent neuroinflammatory response that exerts amyloid ß (Aß) neurotoxic effects, has an effect on endothelial cells, along with the underlying mechanisms. METHODS: In vivo, 5 µL of disease venom was injected into the lateral ventricle of APP/PS1 mice for treatment. Pyroptosis was induced by treating astrocytes with Aß42 in vitro. Small interfering RNA (siRNA) was used to silence caspase-1 and Gasdermin D (GSDMD) mRNA expression. Cell viability was determined using a CCK-8 detection kit. Scanning electron microscopy (SEM), Annexin V/propidium iodide (PI) double staining, RT-qPCR, immunofluorescence, western blotting, and enzyme-linked immunosorbent assay (ELISA) were used to detect cell pyroptosis. The degree of pathological damage to the brain and aortic tissue was assessed by hematoxylin-eosin staining and immunohistochemistry. RESULTS: Aß42 induced astrocyte pyroptosis dependent on the GSDMD/Gasdermin E (GSDME)/Caspase 11/NLRP3 pathway, releasing large amounts of inflammatory factors, such as TNF-α, IL-1α, IL-1ß, and IL-18. Astrocyte pyroptosis caused endothelial cell dysfunction and release of large amounts of vasoconstrictors (ET and vWF). Knockdown of GSDMD reduced astrocyte pyroptosis in the cerebral cortex and hippocampal tissue, decreased the release of inflammatory factors IL-1 ß and IL-18, reduced Aß deposition and tau protein, increased the release of peripheral vasodilator substances (eNOS), and decreased the release of vasoconstrictor substances (ET, vWF), thereby reducing brain tissue damage and vascular injury in APP/PS1 mice. CONCLUSION: Aß42 induced astrocyte pyroptosis, while GSDMD knockout inhibited astrocyte pyroptosis, reduced the release of inflammatory factors, and alleviated brain tissue damage and vascular damage in APP/PS1 mice. Therefore, GSDMD is a novel therapeutic target for Alzheimer's disease.

MiR-590-3p Promotes the Phenotypic Switching of Vascular Smooth Muscle Cells by Targeting Lysyl Oxidase.

ABSTRACT: We investigated the clinical characteristics of patients with acute aortic dissection (AAD) and miR-590-3p levels in serum, tissue, and vascular smooth muscle cells. The effect of miR-590-3p on the vascular smooth muscle cell phenotype was assessed, and the regulation of lysyl oxidase by miR-5903p was determined. C57BL/6 mice were used to investigate the incidence of AAD and effects of miR-5903p on AAD. The miR-590-3p levels were measured in the aortae of mice, and hematoxylin and eosin staining and Masson staining were performed to identify the morphological features of the aorta. Comparative analysis revealed significant differences in clinical characteristics between patients with AAD and healthy control subjects, with most patients with AAD exhibiting concomitant hypertension and nearly 50% having atherosclerosis. Lysyl oxidase was a direct target of miR-590-3p. Lysyl oxidase overexpression inhibited switching of the vascular smooth muscle cell phenotype from contractile to synthetic, but miR-590-3p overexpression significantly reversed this change. In the mouse model, miR-590-3p upregulation increased the incidence of AAD to 93.3%, and its incidence decreased to 13.3% after miR-590-3p inhibition. Hematoxylin and eosin and Masson staining revealed that the miR-590-3p agomiR group had a greater loss of the contractile phenotype in the dissected aortic wall and an increased number of muscle fibers in the aortic wall, which contributed to thickening of the aortic wall and the formation of a false lumen in aortic dissection. miR-590-3p might be pivotal in the pathogenesis of AAD. Thus, targeting miR-590-3p or its downstream pathways could represent a therapeutic approach for AAD.

Hematoxylin and Eosin staining of PhenoCycler® Fusion flow cell slides.

Multiplexed Imaging technologies are powerful techniques that enable ultrahigh-plex spatial phenotyping of whole tissue sections at single cell spatial resolution. Co-Detection by Indexing (CODEX) multiplexing can detect up to 100 proteins using cyclic detection of DNA conjugated antibodies applied to tissue sections. However, it is necessary to correlate multiplexed fluorescent (mIF) spatial images with Hematoxylin and Eosin (H&E) stained sections post analysis. To effectively correlate mIF spatial images with H&E morphology, an (H&E) staining protocol was developed that is directly applied to the CODEX Fusion flow-cell slide after analysis allowing for direct H&E correlation and annotation with mIF images.

Lupenone improves type 2 diabetic nephropathy by regulating NF-κB pathway-mediated inflammation and TGF-ß1/Smad/CTGF-associated fibrosis.

BACKGROUND: Type 2 diabetic nephropathy is a common diabetic complication and the main cause of death in patients with diabetes. Research has aimed to find an ideal drug with minimal side effects for treating this disease. Banana peel has been shown to be anti-diabetic, with lupenone isolated from banana peel exhibiting antidiabetic and anti-inflammatory activities; However, the effects of lupenone on type 2 diabetic nephropathy are largely unknown. PURPOSE: This study aimed to investigate the ameliorative effect of lupenone on type 2 diabetic nephropathy, and its mechanism from both anti-inflammatory and anti-fibrotic perspectives. METHODS: Spontaneous type 2 diabetic nephropathy db/db mouse models were given three levels of lupenone (24 or 12 or 6 mg/kg/d) via intragastric administration for six weeks, and irbesartan treatment was used for the positive control group. We explored the effects and mechanism of lupenone action using enzyme-linked immunosorbent assay, automatic biochemical analyzer, hematoxylin-eosin and Masson staining, real time-PCR, and western blotting. Concurrently, a high-sugar and high-fat diet combined with a low-dose streptozotocin-induced type 2 diabetic nephropathy rat model was used for confirmatory research. RESULTS: Lupenone administration maintained the fasting blood glucose; reduced glycosylated hemoglobin, insulin, and 24 h proteinuria levels; and markedly regulated changes in biochemical indicators associated with kidney injury in serum and urine (including 24 h proteinuria, micro-albumin, N-acetyl-ß-d-glucosaminidase, α1-micro-globulin, creatinine, urea nitrogen, uric acid, total protein, and albumin) of type 2 diabetic nephropathy mice and rats. Hematoxylin-eosin and Masson staining as well as molecular biology tests revealed that inflammation and fibrosis are the two key processes affected by lupenone treatment. Lupenone protected type 2 diabetic nephropathy kidneys by regulating the NF-κB-mediated inflammatory response and TGF-ß1/Smad/CTGF pathway-associated fibrosis. CONCLUSION: Lupenone has potential as an innovative drug for preventing and treating diabetic nephropathy. Additionally, it has great value for the utilization of banana peel resources.

Salidroside Inhibits Ganglion Cell Apoptosis by Suppressing the Müller Cell Inflammatory Response in Diabetic Retinopathy.

PURPOSE: This study aimed to investigate the role of salidroside (SAL) in the cellular communication between Müller cells and retinal ganglion cells in diabetic mice. METHODS: The diabetes mellitus (DM) animal models were established by the intraperitoneal injection of streptozotocin and treatment with SAL via gavage or by the injection of IL-22BP into the vitreous cavity. Immunohistochemistry was used to measure the expression of the glial fibrillary acidic protein in Müller cells. The expression of IL-22 and IL-22Rα1 in retinal tissues was assessed by immunofluorescence. Western blotting was used to measure the expression of inflammatory and apoptosis-related proteins. Hematoxylin-eosin staining, TUNEL staining, and flow cytometry were used to analyze the apoptosis of retinal ganglion cells. The effect of cellular interactions was explored by Transwell assays. RESULTS: Western blotting showed that glial fibrillary acidic protein, IL-22 protein expression was significantly upregulated in the DM animal models compared with the control mice. Immunofluorescence showed that IL-22 was highly expressed in Müller cells and IL-22Rα1 was expressed in ganglion cells in the retina of DM mice. Hematoxylin-eosin and TUNEL staining results showed an increase in the number of ganglion cells apoptotic in DM. However, SAL reversed these phenomena. Meanwhile, after coculture with Müller cells, Western blotting suggested that ganglion cells secreted p-STAT3, and c-caspase3 protein expression was increased. More interestingly, the treatment of IL-22BP and SAL inhibited the expression of the p-STAT3 and c-caspase3 proteins. Flow cytometry indicates that compared with the control group, the apoptosis rate of ganglion cells was increased in the high glucose group, while the apoptosis rate of cells in the recombinant IL-22 protein group was significantly increased, while the SAL inhibited ganglion cells apoptosis. CONCLUSION: SAL inhibits the apoptosis of retinal ganglion cells via the IL-22/STAT3 pathway in Müller cells.

Systemic Diclofenac Sodium Reduces Postoperative rhBMP-2 Induced Neuroinflammation: A Rodent Model Study.

STUDY DESIGN: This is a basic science, animal research study. OBJECTIVE: This study aims to explore, in rodent models, the effectiveness of systemic nonsteroidal anti-inflammatory drugs in reducing recombinant human bone morphogenetic protein-2 (rhBMP-2) induced neuroinflammation. SUMMARY OF BACKGROUND DATA: rhBMP-2 is increasingly used to augment fusion in lumbar interbody fusion surgeries, although it can cause complications including postoperative radiculitis. MATERIALS AND METHODS: Eighteen 8-week-old Sprague-Dawley rats underwent Hargreaves testing to measure the baseline thermal withdrawal threshold before undergoing surgical intervention. The L5 nerve root was exposed and wrapped with an Absorbable Collagen Sponge containing rhBMP-2. Rats were randomized into 3 groups: (1) Low dose (LD), (2) high dose (HD) diclofenac sodium, and (3) saline, receiving daily injection treatment. Hargreaves testing was performed postoperatively on days 5 and 7. Seroma volumes were measured by aspiration and the nerve root was then harvested for hematoxylin and eosin, immunohistochemistry, Luxol Fast Blue staining, and real-time quantitative polymerase chain reaction. The Student t test was used to evaluate the statistical significance among groups. RESULTS: The intervention groups showed reduced seroma volume, and a general reduction of inflammatory markers (MMP12, MAPK6, GFAP, CD68, and IL18) compared with controls, with the reduction in MMP12 being statistically significant ( P = 0.02). Hematoxylin and eosin and immunohistochemistry of the nerve roots showed the highest macrophage density in the saline controls and the lowest in the HD group. Luxol Fast Blue staining showed the greatest extent of demyelination in the LD and saline groups. Lastly, Hargreaves testing, a functional measure of neuroinflammation, of the HD group demonstrated a minimal change in thermal withdrawal latency. In contrast, the thermal withdrawal latency of the LD and saline groups showed a statistically significant decrease of 35.2% and 28.0%, respectively ( P < 0.05). CONCLUSION: This is the first proof-of-concept study indicating that diclofenac sodium is effective in alleviating rhBMP-2-induced neuroinflammation. This can potentially impact the clinical management of rhBMP-2-induced radiculitis. It also presents a viable rodent model for evaluating the effectiveness of analgesics in reducing rhBMP-2-induced inflammation.

Selenium deficiency causes oxidative stress and activates inflammation, apoptosis, and necroptosis in the intestine of weaned calves.

Selenium performs a variety of biological functions in organisms, including antioxidant and anti-inflammatory effects. This study investigated how selenium deficiency affects weaned calves' intestines. According to Inductively coupled plasma mass spectrometry (ICP-MS) analysis of intestinal selenium concentrations in calves, the Se-D group had a significantly lower concentration of selenium. Hematoxylin-eosin staining showed that the intestinal epithelial cells were detached, the goblet cells were lost, and the intestinal villi were fragmented and loosely arranged in the Se-D group, along with hyperemia and inflammatory infiltration. Of the 22 selenoprotein genes, 9 were downregulated in response to selenium deficiency in Reverse transcription-PCR (RT-PCR), whereas 6 genes were upregulated. In the Se-D group, oxidative stress was detected by measuring redox levels in the intestines. Furthermore, TdT-mediated dUTP Nick-End Labeling (TUNEL) staining, RT-PCR, and Western blotting (WB) results indicated that both intrinsic and extrinsic apoptosis pathways are activated in the intestine during selenium deficiency. Selenium deficiency also induced necroptosis in the intestine through upregulation of MLKL, RIPK1, and RIPK3 mRNA levels. In addition, according to hematoxylin-eosin staining and ELISA, selenium-deficient calves had severe inflammation in their intestines. As a result of RT-PCR and WB analyses, we found that selenium deficiency was associated with nuclear factor kappa-B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways. Our study suggested that weaned calves' intestines are affected by selenium deficiency, which causes oxidative stress, inflammation, apoptosis, and necroptosis.

In silico investigation of binding propensity of hematoxylin derivative and damnacanthal for their potential inhibitory effect on HIV-1 Vpr from different subtypes.

HIV-1, the causative agent of AIDS leads to many deaths worldwide though few options are available as therapeutics. To deal with the continuous mutation in the virus genome, requirement of new drugs is always there. Subtype variation plays a crucial role in case of HIV-1 therapeutics development. In this study, we want to investigate some pre examined molecules that can be effective for HIV-1 VPR. Inhibition of several protein-protein interactions with the small molecules will lead to identify some molecules as therapeutics other than the conventional drugs. We retrieved the sequences of different subtypes from the database and representative sequences were identified. Representative structures were modelled and validated using MD simulations. Forty molecules, showing anti Vpr activity in vitro were identified from literature survey and those were docked with each subtype representative structures. Two molecules a stable Hematoxylin Derivative (SHD) and Damnacanthal (D3), these were shown to be bind more effectively for all the subtypes. The stability of the protein and those two small molecule complexes were identified again with MD simulation followed by the binding energy calculation. Thus, these molecules can be thought as any option other than the conventional drug targeting HIV-1 Vpr.Communicated by Ramaswamy H. Sarma.

Danhong injection alleviates cerebral ischemia-reperfusion injury by inhibiting autophagy through miRNA-132-3p/ATG12 signal axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Danhong injection (DHI) is a renowned traditional Chinese medicine often used clinically to treat cardiovascular and cerebrovascular diseases. Studies have shown that DHI can significantly alter microRNA (miRNA) expression in the brain tissue. Therefore, exploring specific miRNAs' regulatory mechanisms during treatment with DHI is essential. AIM OF THE STUDY: To investigate DHI's regulatory mechanism on cerebral autophagy in rats with cerebral ischemia-reperfusion injury (CIRI). MATERIAL AND METHODS: Rats were randomly divided into the sham, middle cerebral artery occlusion (MCAO) model, and DHI-treatment groups. The extent of brain damage was evaluated using triphenyl tetrazolium chloride and hematoxylin-eosin staining. Hippocampal cell autophagy was observed using transmission electron microscopy. Autophagy-related proteins were analyzed using western blotting. Differentially expressed miRNAs were screened using high-throughput and real-time quantitative reverse transcription PCR. The relationship between miR-132-3p and ATG12 was confirmed using a dual-luciferase assay. The miR-132-3p mimics and inhibitors were transfected into PC12 cells subjected to oxygen-glucose deprivation (OGD) in vitro and MCAO model rats in vivo. RESULTS: DHI significantly altered the miRNA expression profile in rat brain tissues. The pathological changes in the brain tissues were improved, and the autophagic hippocampal cell vehicles were significantly reduced after DHI treatment. miRNA-132-3p, one of the miRNAs with a significantly different expression, was screened. Kyoto Encyclopedia of Genes and Genomes signal pathway analysis showed that its target genes were closely related to autophagy. Western blotting revealed that the p-PI3K, p-AKT, and mTOR expression increased significantly; AMPK, ULK1, ATG12, ATG16L1, and LC3II/I were downregulated in the DHI group. Dual-luciferase reporter gene experiments showed that miRNA-132-3p could target the ATG12 3'-UTR region directly. In vitro, miRNA-132-3p had a protective effect on OGD/R-induced oxidative stress injury in PC12 cells, improving cell viability, and affecting the expression of autophagy pathway-related proteins. In vivo transfection experiments showed that miR-132-3p could regulate ATG12 expression in CIRI rats' lateral brain tissue, affecting the autophagy signaling pathway. miR-132-3p overexpression reduces CIRI-induced autophagy and protects neurons. CONCLUSION: This study showed that DHI inhibits neuronal autophagy after cerebral ischemia-reperfusion. This may have resulted from miR-132-3p targeting ATG12 and regulating the autophagy signaling pathway protein expression.

23-O-acetylshengmanol-3-O-α-L-arabinoside alleviates lipopolysaccharide-induced acute lung injury through inhibiting IκB/NF-κB and MAPK/AP-1 signaling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Cimicifuga foetida L. is a well-established traditional Chinese medicine with heat-clearing and detoxifying effects and has good therapeutic effect on oral mucosal ulcer and pharyngitis. The rhizome of this herb is rich in triterpenoid glycosides, including 23-O-acetylshengmanol-3-o-α-L-arabinoside (DA). AIM OF THE STUDY: Whether and how DA attenuates acute lung injury (ALI) are unclear. Accordingly, we focused on its anti-inflammatory effects and underlying molecular mechanisms in lipopolysaccharide (LPS)-stimulated ALI mice and RAW264.7 cells. MATERIALS AND METHODS: The model of ALI mice was established by exposed intratracheal instillation of LPS. Lung pathological changes were evaluated by hematoxylin and eosin staining. Pulmonary function was assessed by whole-body plethysmography. Total protein content in bronchoalveolar lavage fluid (BALF) was detected by bicinchoninic acid method. Wet/dry lung ratio was used to evaluate the degree of pulmonary edema in mice. The levels of pro-inflammatory mediators were measured using enzyme-linked immunosorbent assay. The relative expression of pro-inflammatory gene mRNA was examined by RT-qPCR. The expression of inflammatory-related proteins was detected by Western blot. RAW264.7 cells were used to test the anti-inflammatory effects of DA in vitro. Cytotoxicity was assessed using a MTT assay. Nitric oxide production was measured by Griess assay. The production and expression of inflammatory mediators and the protein levels of inflammatory signaling molecules in the NF-κB and MAPK pathways were measured. Furthermore, immunofluorescence staining was used to analyze the expression of p-IκBα, p-ERK, and p-p38 in lung macrophages and the nuclear translocation of NF-κB p65 and AP-1 in cells. RESULTS: DA evidently alleviated histopathological changes and ameliorated pulmonary edema. Moreover, DA could reduce excessive inflammatory reaction in lung tissue as manifested by the reduction of proinflammatory mediators (IL-1ß, IL-6, TNF-α, MCP-1, iNOS, and COX-2) in BALF, serum, and lung tissues. Further, DA inhibited the activation of the NLRP3/caspase-1 pathway in the lung. DA reduced the production and expression of the proinflammatory mediators above in RAW264.7 cells. Mechanistically, DA remarkably blocked the nuclear translocation of NF-κB p65, suppressed IκBα phosphorylation, and markedly reduced the nuclear translocation of AP-1 and the phosphorylation of ERK and p38. CONCLUSIONS: The findings demonstrated that DA exerts anti-inflammatory effects in LPS-stimulated ALI mice and macrophages by downregulating the NLRP3/caspase-1 signaling pathway in lung tissue and the IκB/NF-κB and MAPKs/AP-1 pathways in macrophages, suggesting that DA may be promising in ALI treatment.

Saffron essential oil ameliorates CUMS-induced depression-like behavior in mice via the MAPK-CREB1-BDNF signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Saffron, the dried stigma of Crocus sativus L., has a long history of use in the treatment of depression in traditional Chinese medicine and Islamic medicine. The unique aroma of saffron, primarily derived from its volatile oil, has been widely used by folk to mitigate anxiety and depression via sniffing because the aroma of saffron has a pleasant and invigorating effect. AIM OF THE STUDY: This study aimed to investigate the antidepressant effect and the underlying mechanism of saffron essential oil (SEO) in mice exposed to chronic unpredictable mild stress (CUMS). MATERIALS AND METHODS: In this study, compounds of SEO were identified using gas chromatography-mass spectrometry analysis, while network pharmacology was used to predict potential active compounds, antidepressant targets, and related signaling pathways of SEO. The CUMS depression model was further used to explore the therapeutic effect and possible mechanism of SEO. During the modeling period, mice were regularly administered fluoxetine (3.6 mg/kg, i.g.) or diluted SEO (2%, 4%, and 6% SEO, inhalation). The antidepressant and neuroprotective effects of SEO were evaluated by behavior tests (the open field test, the sucrose preference test, the tail suspension test, and the forced swimming test), hematoxylin-eosin staining, and Nissl staining. The enzyme-linked immunosorbent assay kits were used to measure dopamine (DA), 5-serotonin (5-HT), brain-derived neurotrophic factor (BDNF), and γ-aminobutyric acid (GABA) levels in serum. The relative abundance of Raf1, MEK1, P-ERK1/2/ERK1/2, P-CREB1/CREB1, BDNF, and P-Trk B/Trk B in the hippocampus was determined using western blot (WB). RESULTS: According to the network pharmacology analysis, seven active SEO compounds mediated 113 targets related to depression treatment, most of which were enriched in the 5-HT synapse, calcium signaling pathway, and cAMP signaling pathway. In vivo experiments indicated that fluoxetine and SEO improved depression-like behaviors in depressed mice. The levels of 5-HT, DA, BDNF, and GABA in serum increased significantly. Histopathological examinations revealed that fluoxetine and SEO ameliorated neuronal damage in the hippocampus. WB analysis showed that the relative expressions of Raf1, MEK1, P-ERK1/2/ERK1/2, P-CREB1/CREB1, BDNF, and P-Trk B/Trk B were significantly higher in the fluoxetine and SEO groups than in the CUMS group. CONCLUSION: Overall, these findings suggest that SEO significantly alleviates the depressive symptoms in CUMS exposed mice and partially restores hippocampal neuronal damage. Meanwhile, the best efficacy was observed in 4% SEO. Furthermore, the antidepressant mechanism of SEO is primarily dependent on the regulation of the MAPK-CREB1-BDNF signaling pathway.

Fufang-zhenzhu-tiaozhi formula protects islet against injury and promotes ß cell regeneration in diabetic mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Fufang-zhenzhu-tiaozhi formula (FTZ) is a patented preparation of traditional Chinese medicine that has been used to treat hyperglycemia and hyperlipidemia in the clinic for almost 10 years. Our previous study had demonstrated that FTZ can protect islet ß cell injury in vitro. However, the efficacy of FTZ on ß cell regeneration in vivo and the involved anti-diabetic mechanism remains unknown. AIM OF THE STUDY: We aim to investigate the effects of FTZ as a good remedy for islet protection and ß cell regeneration, and to reveal the underlying mechanism. MATERIALS AND METHODS: C57BL/6 mice were fed with high-fat diet for 3 weeks and then intraperitoneally injected with streptozotocin (90 mg/kg/d × 1 d) to establish type 2 diabetes (T2D) models. Mice in each group were divided into three batches that sacrificed after 3, 7 and 28 days of FTZ administration. Body weight, blood glucose, and oral glucose tolerance test were measured at indicated time points. Fasting insulin was determined by enzyme-linked immunosorbent assay (ELISA) kit. Neonatal ß cell was assessed by insulin & PCNA double immunofluorescence staining, and the underlying mechanisms related to ß cell regeneration were further performed by hematoxylin-eosin staining, insulin & glucagon double immunofluorescence staining and Western blot. RESULTS: FTZ and metformin can significantly help with the symptoms of DM, such as alleviating weight loss, reducing blood glucose, improving the level of insulin in vivo, and relieving insulin resistance, suggesting FTZ and metformin treatment maintained the normal morphological function of islet. Notably, ß cell regeneration, which is indicated by insulin and PCNA double-positive cells, was promoted by FTZ, whereas few neonatal ß cells were observed in metformin group. Hematoxylin-eosin staining, and its quantification results showed that FTZ effectively prevented the invasion of inflammatory cells into the islets in diabetic mice. Most ß cells in the islets of diabetic model mice were devoid, and the islets were almost all α cells, while the diabetic mice administered FTZ could still maintain about half of the ß cells in the islet. Furthermore, FTZ upregulated the expression of critical transcription factors during ß cell development and maturation (such as PDX-1, MAFA and NGN3) in diabetic mice. CONCLUSIONS: FTZ can alleviate diabetes symptoms and promote ß cell regeneration in diabetic mice. Moreover, FTZ promotes ß cell regeneration by preserving islet (resisting inflammatory cells invading islets), maintaining the number of ß cells in islets, and increasing the expression of PDX-1, MAFA and NGN3.

The impact of IGF-1 on alveolar bone remodeling and BMP-2 expression in orthodontic tooth movement in diabetic rats.

BACKGROUND: Orthodontic tooth movement is linked to alveolar bone reconstruction. OBJECTIVES: As a regulator of cell proliferation, insulin-like growth factor 1 (IGF-1) plays an important role in osteoporotic fracture healing. This study aims to investigate the effect of IGF-1 on alveolar bone remodeling in diabetic rats. MATERIAL AND METHODS: Sprague Dawley (SD) rats were randomly divided into 3 groups, including a control group, a model group established with streptozotocin (STZ) injection to prepare the diabetic rats (type 1 diabetes), and an IGF-1 group of diabetic rats receiving daily intraperitoneal injections of 1.0 mg/kg IGF-1. Nickel-titanium coil springs were used to pull the first molar forward to establish the model. The maxillary first to third molars and the surrounding alveolar bone were collected to measure tooth movement distance. Hematoxylin and eosin (H&E) staining was applied to detect the pathological changes in the periodontal tissue. Real-time polymerase chain reaction (PCR) and western blot were adopted to measure bone morphogenetic protein 2 (BMP-2) mRNA and protein expression. Enzyme-linked immunosorbent assays (ELISAs) were used to measure interleukin-1α (IL-1α) levels in the serum. RESULTS: The tooth movement distance was significantly decreased, BMP-2 expression was downregulated, and IL-lα levels were enhanced in the model group compared to the control group (p < 0.05). However, the tooth movement distance was increased, BMP-2 expression was increased, and IL-lα levels were reduced in the IGF-1 group compared to the model group (p < 0.05). Hematoxylin and eosin staining showed that alveolar bone destruction was attenuated in the IGF-1 group, while the new bone was not active in the model group. CONCLUSIONS: Diabetes can damage alveolar bone remodeling in orthodontic tooth movement. The IGF-1 promotes alveolar bone remodeling by inhibiting inflammation and upregulating BMP-2 expression.

Xianglian Pill attenuates ulcerative colitis through TLR4/MyD88/NF-κB signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Xianglian Pill (XLP) is a classical Chinese medicine prescription applied for controlling ulcerative colitis (UC). Whereas, the underlying mechanism remains unclear. AIM OF THE STUDY: The present work was aimed to investigate the mechanism of XLP in dextran sulfate sodium (DSS)-induced UC via the Toll Like Receptor 4 (TLR4)/Myeloid Differentiation factor 88 (MyD88)/Nuclear Factor kappa-B (NF-κB) signaling in mice. MATERIALS AND METHODS: The major components of XLP were detected by high-performance liquid chromatography-diode array detection (HPLC-DAD). The ulcerative colitis model was induced by DSS in mice. 5-Amino Salicylic Acid (5-ASA) group and XLP group were intragastrically treated. Disease activity index (DAI) and colon length were monitored and hematoxylin-eosin (HE) staining was conducted. Gasdermin D (GSDMD)-N and TLR4 expressions in colon tissues were visualized by immunofluorescence. TLR4 mRNA was measured by Real Time Quantitative PCR (RT-qPCR). The expressions of NOD-like receptor thermal protein domain associated protein 3 (NLRP3), active-caspase-1, GSDMD-N, TLR4, MYD88, NF-κB, p-NF-κB, and the ubiquitination of TLR4 in colon tissues were detected by Western blot. Myeloperoxidase (MPO) enzyme activity was examined and serum inflammatory factors Interleukin (IL)-1ß, IL-6, Tumor Necrosis Factor-α (TNF-α), and IL-18 were determined by Enzyme-linked Immunosorbent Assay (ELISA). TLR4-/- mice were applied for verifying the mechanism of XLP attenuated DSS symptoms. RESULTS: The XLP treatment extended colon length, reduced DAI, and attenuated histopathological alteration in DSS-induced mice. XLP administration suppressed MPO activity and reduced the content of IL-1ß, IL-6, TNF-α and IL-18 in serum. XLP also inhibited the expression levels of GSDMD-N, TLR4, NLRP3, active-caspase-1, MyD88, p-NF-κB/NF-κB in colon tissues of DSS-induced mice. TLR4-/- mice proved that TLR4 was involved in XLP-mediated beneficial effect on DSS-induced ulcerative colitis. CONCLUSIONS: XLP might treat ulcerative colitis by regulating the TLR4/MyD88/NF-κB signaling pathway.

Swimming exercise and nano-l-arginine supplementation improve oxidative capacity and some autophagy-related genes in the soleus muscle of aging rats.

The present research aims to evaluate the effect of swimming exercise and chitosan-coated l-arginine on mitochondrial oxidation, BCL2 Interacting Protein 3 (Bnip3), NIP-like protein × (Nix), B-cell lymphoma-extra-large (Bcl-xL) and autophagy-related protein light chain 3(LC3) expression in soleus muscle of aging rats. In this experimental research, 25 male Wistar rats were assigned into five groups randomly: young, old, old + Nano l-arginine (Nano L-a), old + exercise (Ex), and old + Nano l-arginine (Nano L-a) + exercise (Ex) (n = 5 in each). They performed a swimming exercise program five days a week for six weeks. To determine the relative strength for rats before and after performing these interventions, the 1repetition maximum (1RM) test was done as a pre and post-test. The exercise program started with 20 min and after four sessions, gradually increased to 60 min and this time was maintained until the completion of the training period. l-arginine coated with chitosan nanoparticles was given to the rats in the l-arginine-supplemented group via gavage at a dosage of 500 mg/kg/day, five days a week, for six weeks. Additionally, the rats in all groups were fed a normal diet (2.87 kcal/g and 15 % energy from fat). Upon the completion of the protocol implementation, the rats were sacrificed and the soleus muscle was fixed and frozen to determine hematoxylin and eosin (H&E) staining, immunohistochemistry (IHC), gene expression analysis, levels of reactive oxygen species (ROS), and total antioxidant capacity (TAC). The results from the present research indicated that swimming exercise and Nano l-arginine improve the strength and histology of muscle tissue in old rats (p < 0.05). Aging significantly increased the expression of Nix and Bnip3 (p < 0.05) and reduced the Bcl-xL gene expression (p < 0.05). The expression of LC3 protein also increased with aging (p < 0.05). Therapeutic interventions, such as combined treatment (old + Nano L-a + Ex) for old animals, reduced the amount of this protein in soleus muscle (p < 0.05). The ROS values also showed a significant reduction only in the old + Nano L-a + Ex group compared to the old group. Moreover, TAC values show a significant decrease in the old and old + Ex groups in comparison to the young group. The use of arginine supplement, especially in nano form, along with swimming exercise seems to reduce the oxidative damage to the elderly muscle tissue, which has a positive effect on the structure and function of the soleus muscle. Since these interventions only had a significant effect on LC3 protein, further studies with more diverse measurement methods for autophagy are suggested.

Ameliorative Effect of Imperatorin on Dermatophagoides pteronyssinus-Induced Allergic Asthma by Suppressing the Th2 Response in Mice.

Imperatorin is a furanocoumarin derivative and an effective ingredient in several Chinese medicinal herbs. It has favorable expectorant, analgesic, and anti-inflammatory effects. In this study, we investigated whether imperatorin has protective effects against Dermatophagoides pteronyssinus (Der p)-induced asthma in mice. Lung and bronchial tissues were histopathologically examined through hematoxylin-eosin staining. The concentrations of immunoglobin E (IgE), IgG1, IgG2a in serum and those of T helper 1 (Th1) and two cytokines and eosinophil-activated chemokines in bronchoalveolar lavage fluid (BALF) were detected using an enzyme immunoassay. Histological examination revealed that imperatorin reduced inflammatory cell infiltration, mucus hypersecretion, and endothelial cell hyperplasia. The examination also indicated that imperatorin could reduce the inflammatory cell count in BALF as well as IgE and IgG1 expression in serum, but IgG2a expression was significantly increased. Imperatorin reduced the production of interleukin (IL)-4, IL-5, and IL-13 by Th2, promoted the production of interferon-γ and IL-12 by Th1, and increased the production of IL-10 in bronchoalveolar lavage fluid. These findings suggest that imperatorin has a considerable anti-inflammatory effect on Der p-induced allergic asthma in mice.

Healing Potential of Propolis Extract-Passiflora edulis Seed Oil Emulgel Against Excisional Wound: Biochemical, Histopathological, and Cytokines Level Evidence.

Propolis is rich in natural bioactive compounds, and considering its importance for many skin therapies, emulgel was prepared. This study examines how a propolis extract (PE) and Passiflora edulis seed (PS) oil emulgel affect rat deep skin wound healing. Based on preset criteria of maximum drug content and optimum drug permeation through the stratum corneum along with drug retention in the skin layers, an optimized emulgel formula based on Box-Behnken factorial design was prepared and used for subsequent in vitro and in vivo evaluations. In vivo wound-healing activities of emulgel and control treatments were investigated in a rat model. The optimized emulgel formula exhibited superior healing activity compared with plain PE suspension-treated rats on day 14 of wounding. Histopathological investigations of hematoxylin and eosin and Masson's Trichrome-stained skin sections supported this effect. Emulgel promotes cutaneous wound healing through a variety of mechanisms, including anti-inflammatory through modulation of cytokines tumor necrosis factor-α, interleukin (IL)-1ß, and IL-6 production, and promotion of collagen fiber formation, all of which contribute to tissue remodeling. Furthermore, when compared with propolis suspension, emulgel showed significant antioxidant and anti-inflammatory effects. Emulgel significantly increased the skin's hydroxyproline level, antioxidant potential, wound contraction, increased penetration, and localized propolis deposition across the skin. Incorporation of PS oil into the emulgel accelerates the tissue regeneration process. The findings suggest that 5% propolis emulgel could be used as an alternative to treat wounds.

Histometric and morphological damage caused by Serratia marcescens to the tick Rhipicephalus microplus (Acari: Ixodidae).

Rhipicephalus microplus tick is the ectoparasite causing the greatest economic losses in the livestock industry. Multi-resistance in ticks is increasing, generating the inefficiency of traditional ixodicides, for which biological control has been proposed as an alternative. In this work, we analyze the histomorphological damage caused by the bacterial strain EC-35 on Rhipicephalus microplus. The ixodicidal effect of EC-35 total protein was evaluated on larval or adult ticks comparing with the commercial ixodicide coumaphos 0.02% as a control. Female ticks were processed using the paraffin-embedding technique and stained with hematoxylin-eosin. Also, the pathogenicity of EC-35 was evaluated by capillary feeding and coelom inoculation tests. The identification of the bacterium was performed using the molecular markers 16S RNA and rpoB, by PCR and sequencing technique, and the evolutionary distance was analyzed by Bayesian phylogenetic inference. No differences were observed in the perimeter and area of larvae treated with EC-35 or Coumaphos. The thickness of the integument decreased a 65% with the EC-35 treatment (6.01 ± 0.6 µm) and of 30% in coumaphos (12.04 ± 1.2 µm) in larvae compared with the control group (18.41 ± 2 µm), while no difference was found in adult ticks. The capillary feeding test and coelom inoculation with EC-35 showed an inhibition of reproductive potential of 99.8 ± 7 and an oviposition Inhibition 97 ± 3.02%. The EC-35 strain was genetically related to Serratia marcescens, concluding that these bacteria caused high mortality, oviposition Inhibition, and integument thinning and drastic loss of histoarchitecture in R. microplus tick larvae.

Zinc Oxide Particles Can Cause Ovarian Toxicity by Oxidative Stress in Female Mice Model.

Introduction: Zinc oxide nanoparticles (ZnO NPs) participate in all aspects of our lives, but with their wide application, more and more disadvantages are exposed. The goal of this study was to investigate the toxicity of ZnO NPs in female mice ovaries and explore its potential mechanism. Methods: In this study, adult female mice were orally exposed to 0, 100, 200, and 400 mg/kg ZnO NPs for 7 days. We explored the underlying mechanisms via the intraperitoneal injection of N-acetyl-cysteine (NAC), an inhibitor of oxidative stress, and salubrinal (Sal), an inhibitor of endoplasmic reticulum (ER) stress. Results: The results indicated that serum estradiol and progesterone levels declined greatly with increasing ZnO NPs dosage. Hematoxylin and eosin (HE) staining revealed increased atretic follicles and exfoliated follicular granulosa cells. Moreover, at the transcriptional level, antioxidant-related genes such as Keap1 and Nrf2, and ER stress-related genes PERK, eIF2α, and ATF4 were markedly upregulated. In addition, the expression of Caspase12, Caspase9, and Caspase3, which are genes related to apoptosis, was also upregulated in all ZnO NPs treatment groups. Serum malondialdehyde (MDA) content was remarkably up-regulated, whereas superoxide dismutase (SOD) activity was down-regulated. The 400 mg/kg ZnO NPs treatment group suffered the most substantial harm. However, ovarian damage was repaired when NAC and Sal were added to this group. Conclusion: ZnO NPs had toxic effects on the ovary of female mice, which were due to oxidative stress, ER stress, and the eventual activation of apoptosis.

Inhibition of inflammasome activation via sphingolipid pathway in acute lung injury by Huanglian Jiedu decoction: An integrative pharmacology approach.

BACKGROUND: Acute lung injury (ALI) is a serious health issue which causes significant morbidity and mortality. Inflammation is an important factor in the pathogenesis of ALI. Even though ALI has been successfully managed using a traditiomal Chinese medicine (TCM), Huanglian Jiedu Decoction (HLD), its mechanism of action remains unknown. PURPOSE: This study explored the therapeutic potential of HLD in lipopolysaccharide (LPS)-induced ALI rats by utilizing integrative pharmacology. METHODS: Here, the therapeutic efficacy of HLD was evaluated using lung wet/dry weight ratio (W/D), myeloperoxide (MPO) activity, and levels of tumor necrosis factor (TNF-α), interleukin (IL)-1ß and IL-6. Network pharmacology predictd the active components of HLD in ALI. Lung tissues were subjected to perform Hematoxylin-eosin (H&E) staining, metabolomics, and transcriptomics. The acid ceramidase (ASAH1) inhibitor, carmofur, was employedto suppress the sphingolipid signaling pathway. RESULTS: HLD reduced pulmonary edema and vascular permeability, and suppressed the levels of TNF-α, IL-6, and IL-1ß in lung tissue, Bronchoalveolar lavage fluid (BALF), and serum. Network pharmacology combined with transcriptomics and metabolomics showed that sphingolipid signaling was the main regulatory pathway for HLD to ameliorate ALI, as confirmed by immunohistochemical analysis. Then, we reverse verified that the sphingolipid signaling pathway was the main pathway involed in ALI. Finally, berberine, baicalein, obacunone, and geniposide were docked with acid ceramidase to further explore the mechanisms of interaction between the compound and protein. CONCLUSION: HLD does have a better therapeutic effect on ALI, and its molecular mechanism is better elucidated from the whole, which is to balance lipid metabolism, energy metabolism and amino acid metabolism, and inhibit NLRP3 inflammasome activation by regulating the sphingolipid pathway. Therefore, HLD and its active components can be used to develop new therapies for ALI and provide a new model for exploring complex TCM systems for treating ALI.