Characterization of sciatic nerve myelin sheath during development in C57BL/6 mice.

Myelin sheath plays important roles in information conduction and nerve injury repair in the peripheral nerve system (PNS). Enhancing comprehension of the structure and components of the myelin sheath in the PNS during development would contribute to a more comprehensive understanding of the developmental and regenerative processes. In this research, the structure of sciatic nerve myelin sheath in C57BL/6 mice from embryonic day 14 (E14) to postnatal 12 months (12M) was observed with transmission electron microscopy. Myelin structure appeared in the sciatic nerve as early as E14, and the number and thickness of myelin lamellar gradually increased with the development until 12M. Transcriptome analysis was performed to show the expressions of myelin-associated genes and transcriptional factors involved in myelin formation. The genes encoding myelin proteins (Mag, Pmp22, Mpz, Mbp, Cnp and Prx) showed the same expression pattern, peaking at postnatal day 7 (P7) and P28 after birth, whereas the negative regulators of myelination (c-Jun, Tgfb1, Tnc, Cyr61, Ngf, Egr1, Hgf and Bcl11a) showed an opposite expression pattern. In addition, the expression of myelin-associated proteins and transcriptional factors was measured by Western blot and immunofluorescence staining. The protein expressions of MAG, PMP22, MPZ, CNPase and PRX increased from E20 to P14. The key transcriptional factor c-Jun co-localized with the Schwann cells Marker S100ß and decreased after birth, whereas Krox20/Egr2 increased during development. Our data characterized the structure and components of myelin sheath during the early developmental stages, providing insights for further understanding of PNS development.

The effect of post-abortion care (PAC) on anxiety in women with spontaneous abortion based on MicroRNA-21 expression, cortisol level, and Fordyce happiness pattern.

Abortion is one of the most common complications in pregnancy, and the cause of its occurrence in many cases remains unknown. The high prevalence and consequences of anxiety in women with spontaneous abortion could highlight the importance and role of post-abortion care (PAC). Detection and identification of biomarkers related to abortion and anxiety can effectively diagnose and prevent complications. Among the known biomarkers, microRNAs and the cortisol level have high potential. Therefore, the present study evaluated the effect of post-abortion care (PAC) on anxiety in women with spontaneous abortion based on MicroRNA-21 expression, cortisol level, and Fordyce happiness pattern. In this randomized clinical trial, 72 women with spontaneous abortion were studied and randomly divided into two groups of intervention (n = 36) and control (n = 36). Data were collected through a demographic questionnaire and HADS. To assess PAC, the intervention group was consulted in 8 sessions of 60 minutes in the first 72 hours after abortion. Meetings were held twice a week for four weeks. Both groups were followed up immediately after and one month after the intervention. To evaluate biological factors, 4ml of blood sample was obtained from the subjects. Blood cortisol levels were measured by the Cortisol Competitive Human ELISA Kit (Thermo-Fisher, USA), and microRNA-21 evaluation was performed by Real-time PCR technique. Data were analyzed using SPSS16 software. Results showed that before the intervention, there was no significant difference in the mean score of anxiety between the control and intervention groups (P> 0.05); But at the time immediately and one month after the intervention, there was a significant difference in the mean score of anxiety (p <0.001). The results of biological factors evaluation showed that in the intervention group, serum cortisol levels and microRNA-21 expression decreased significantly (p <0.05). In general, PAC based on the happiness pattern can control the anxiety of women with spontaneous abortion. Therefore, it is recommended as an effective and non-invasive intervention in preventing women's psychological problems after spontaneous abortion.

Neuromuscular electrical stimulation effect on mRNA expression of skeletal muscle in elderly.

Considering the increasing number of elderly in the world, this research aimed to investigate the effect of neuromuscular electrical stimulation (NMES) on changes in muscle mRNA abundance of a number of gene targets for improving the balance of the elderly. Twenty-six elderly undertook 30 minutes of quadriceps NMES (50 Hz, current at the limit of tolerance). Vastus lateralis muscle biopsies were obtained at rest immediately before and 24 hours after the intervention. The expression of 384 targeted mRNA transcripts was assessed by Real-time TaqMan PCR. A significant change in expression from baseline was determined using the ΔΔCT method with a false discovery rate (FDR) of <5%. The results showed that the biological functions of upregulated genes included muscle protein turnover, hypertrophy, inflammation, and muscle growth, while downregulated genes included mitochondrial and cell signaling functions. In general conclusion, it can be said that NMES can improve balance in the elderly. Therefore, considering the importance of balance in old people, it is suggested to use this method to improve the balance of the elderly.

[Effect of granules made by new-medicinal parts of Crocus sativus on hyperuricemia in rats and its mechanism].

To explore the effect of the granules made by new-medicinal parts of Crocus sativus(NMPCS) on hyperuricemia(HUA) in rats, the rat model of HUA was established by intramuscular injection of 3% potassium oxonate and intraperitoneal injection of 4% pyrazinamide. The content of serum uric acid was monitored every week for 3 consecutive weeks. After the experiment, the levels of serum uric acid, urine uric acid, serum creatinine, blood urea nitrogen(BUN), and xanthine oxidase(XOD) were determined. The protein and gene expressions of XOD were determined by Western blot method and fluorescence quantitative polymerase chain reaction(qPCR), and the morphological changes in the liver tissue were performed by hematoxylin-eosin(HE) staining. The results showed that as compared with the model group, the levels of serum uric acid in the positive drug group and the low, medium, and high-dose NMPCS groups were lower(P<0.05), the levels of urine uric acid in the high-dose NMPCS group were decreased(P<0.01), and there was no statistical difference in the medium and low-dose NMPCS groups. The levels of BUN in the high and low-dose NMPCS groups were decreased(P<0.05), and the levels of serum creatinine did not change in the administration groups. The positive drug group and the low, medium, and high-dose NMPCS groups significantly reduced the liver damage, with only a few hepatocytes vacuolization and a small number of red blood cells in the central venous area. The nephridial tissue structure was slightly abnormal, with a small number of red blood cell infiltration, and no obvious inflammatory cell infiltration was found in the glomerulus in these groups. No degeneration was found in renal tubular epithelial cells, with mild glomerular and tubular lesions and a small amount of sodium urate deposition and crystallization in the positive drug group and the low, medium, and high-dose NMPCS groups. The relative protein expressions of XOD in the positive drug group and the high dose NMPCS group were decreased(P<0.05), and the relative mRNA expressions of XOD in the positive drug group and the high and low-dose NMPCS groups were decreased as well(P<0.05). The above results show that NMPCS reduces uric acid in rats with HUA by regulating XOD, which provides a certain experimental basis for the development of NMPCS as a new medicine for the treatment of HUA.

[Analysis on transcriptionomic characteristics of Naoxintong Capsules in prevention of post-ischemic inflammation based on RNA-Seq technology].

In this research, high-throughput sequencing was used to investigate the mechanism of Naoxintong Capsules(NXTC) in prevention of post-ischemic inflammation. First, microglia BV-2 inflammatory model was induced by 1.0 µg·mL~(-1) LPS to investigate the effect of intestinal absorption solution of NXTC(NXTCIA) at different concentrations(62.5, 31.25, 15.63, 7.81 µg·mL~(-1)) on LPS-induced BV-2 inflammatory factors in microglia. Then, an RNA-Seq high-throughput sequencing method was performed to identify the differentially expressed mRNAs in microglia BV-2 after pre-treatment with NXTC. GO and KEGG enrichment analysis was used to screen the potential biological processes and related signaling pathways of NXTC in inhibiting inflammation. The results showed that four NXTCIA concentrations could significantly inhibit the release of LPS-induced inflammatory mediators in BV-2 in a dose-dependent manner. Furthermore, high-throughput sequencing results showed that 392 mRNA transcripts were reversed following pre-treatment with NXTC. GO enrichment analysis showed that the transcripts reversed by NXTC were mainly involved in Toll-like receptor signaling pathway, chemokine signaling pathway, and TNF signaling pathway. Taken together, our findings showed that NXTC treatment could provide protective effects against inflammatory response and the mechanism might be related to the regulation of Toll-like receptor signaling pathway, chemokine signaling pathway, and TNF signaling pathway.

Increased Expression of Ubiquitin-Specific Protease 4 Participates in Neuronal Apoptosis After Intracerebral Hemorrhage in Adult Rats.

Ubiquitinating enzymes catalyze protein ubiquitination, a reversible process countered by deubiquitinating enzyme (DUB) action. Ubiquitin-specific protease 4 (USP4) is a member of the ubiquitin-specific protease (USP) family of DUBs that has a role in spliceosome regulation. In the present study, we demonstrated that USP4 may be involved in neuronal apoptosis in the processes of intracerebral hemorrhage (ICH). We obtained a significant up-regulation of USP4 in neurons adjacent to the hematoma following ICH by the results of Western blot, immunohistochemistry, and immunofluorescence. Increasing USP4 level was found to be accompanied by the up-regulation of active caspase-3, γH2AX, Bax, and decreased expression of Bcl-2. In addition, USP4 co-localized well with γH2AX in the nucleus in the ICH model and hemin-induced apoptosis model. Moreover, in vitro study, knocking down USP4 by USP4-specific siRNA in PC12 cells reduced active caspase-3 expression. All these results above suggested that USP4 may be involved in neuronal apoptosis after ICH.

Spatiotemporal Patterns of RING1 Expression after Rat Spinal Cord Injury.

Ring finger protein 1 (RING1) is a RING domain characterized protein belonging to the RING finger family. It is an E3 ubiquitin-protein ligase that mediated monoubiquitination of histone H2A and the core component of PRC1 complex, which is the repressive multiprotein complex of Polycomb group (PcG). Previous studies showed the important tumorigenic role of RING1 via promoting cell proliferation and the crucial function in maintaining transcriptional program stability during development. However, its mechanism for spinal cord injury (SCI) is still unknown. In our research, we established an acute SCI model in adult rats and studied the expression and function profiles of RING1. RING1 protein level detected by western blot peaked at day 3 after trauma and then decreased gradually. Immunohistochemistry showed the increase of RING1 expression displayed in the white matter more obviously than in the gray matter. Furthermore, increased co-expression of RING1 and GFAP confirmed activated astrocytes in injured spinal cord via double immunofluorescence staining. Meanwhile, we also found the co-localization of PCNA, a famous marker of proliferative cells, with RING1 and GFAP, which indicated RING1 might play a role in astrocyte proliferation after SCI. In vitro studies, RING1 protein level in C6 cells increased after LPS challenge and RING1 was required for astrocyte proliferation and activation induced by LPS. In summary, we took a new insight into the function of RING1 in the cellular and molecular mechanism underlying the pathophysiology of SCI.

[Protective effect and mechanism of scutellarin ethyl ester on focal cerebral ischemia induced by ligation of middle cerebral artery in rats].

To observe the protective effect of scutellarin ethyl ester on focal cerebral ischemia injury induced by middle cerebral artery occlusion in rats(MCAOR), and explore its mechanism. Totally 84 male SD rats were randomly divided into seven groups： sham-operated group, model group,positive drug group(niomdipine,12 mg•kg⁻¹), Brevisapin tablets group(48 mg•kg⁻¹), and high, middle and low-dose scutellarin ethyl ester groups(100, 50, 25 mg•kg⁻¹). The MCAOR model was prepared by using thread embolism method to observe the neurological function of rats, the area of cerebral infarction was measured with TTC, and the levels of MDA, SOD and NO in serum were detected with semiautomatic biochemistry analyzer.Ox-LDL and TNF-α cell injury models was established by treating HUVECs with 200 mg•L⁻¹ ox-LDL and 100 µg•L⁻¹ TNF-α,and the levels of MDA, SOD, NO, ET, 6-keto-PGF1α,TXB2, IL-1, IL-6, IL-8, ICAM-1 and PECAM-1 in the cell supernatant were determined. The results showed that scutellarin ethyl ester could effectively improve the neurological function of MCAOR rats, and significantly reduce the area of cerebral infarction. Compared with the model group, activities of SOD and NO in serum increased, while content of MDA decreased. In the cell supernatant, activities of SOD, 6-keto-PGF1α and NO increased, content of IL-1, IL-6, IL-8, ICAM-1, PECAM-1, TXB2, ET and MDA decreased, which indicated that scutellarin ethyl ester has a certain protective effect on focal cerebral ischemia injury induced by middle cerebral artery occlusion in rats, and its mechanism may be related to antioxidative stress, improvement of endothelial function and reduction in inflammatory reaction.

[Simultaneous determination of seven bioactive compounds and pharmacokinetics in rat plasma after oral administration of Yindan Xinnaotong Ruanjiaonang by UPLC-MS/MS].

To estabish ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for simultaneous determination of quercetin(QCT), isorhamnetin(ISR), kaempferol(KMF), ginkgolide A(GA), ginkgolide B(GB), ginkgolide C(GC) and bilobalide(BB) in rat plasma and investigate the pharmacokinetic process of seven compounds after oral administration of Yindan Xinnaotong Ruanjiaonang, The results indicated that all calibrations curves showed good linearity (r≥0.997 1). RSD of intra-day and inter-day precisions were all within 11%. The matrix effects and extraction recovery were in the range of 93.28%-103.6% and 72.43%-95.77% respectively. The peak concentration (Cmax) of QCT, ISR, KMF, GA, GB, GC and BB were (45.02±11.28), (49.90±13.82), (27.85±8.38), (76.31±18.19), (76.54±15.43), (35.35±10.28), (48.70±12.34) µg•L⁻¹, respectively. The peak time (tmax) of seven constituents were (0.33±0.11), (0.50±0.23), (0.33±0.14), (0.75±0.29), (1.0±0.35), (1.5±0.23), (0.75±0.50) h, respectively. UPLC-MS/MS method established in this research was proved to be so rapid and sensitive that it can be applied to the pharmacokinetic study of seven bioactive constituents in Yindan Xinnaotong Ruanjiaonang.

Toll-Interleukin 1 Receptor domain-containing adaptor protein positively regulates BV2 cell M1 polarization.

Microglial activation, including classical (M1) and alternative (M2) activation, plays important roles in the development of several central nervous system disorders and promotes tissue reconstruction. Toll-like receptor (TLR)4 is important for microglial polarization. TIR domain-containing adaptor protein (TIRAP) is an intracellular adaptor protein, which is responsible for the early phase of TLR4 activation. The role of TIRAP in BV2 cell M1 polarization is still unknown. In this study, we showed that TIRAP expression is greatly elevated in lipopolysaccharide (LPS)/interferon (IFN)-γ-treated microglia. TIRAP overexpression promoted BV2 microglial M1 polarization by increasing M1-related marker production (inducible nitric oxide synthase, CD86, interleukin-6, interleukin-1ß and tumour necrosis factor-α). In contrast, TIRAP knockdown prevented M1-related marker production. Mechanistically, TIRAP could interact with TNF Receptor-Associated Factor 6 (TRAF6) to increase M1-related marker production in TIRAP overexpressed and LPS/IFN-γ-treated BV2 cells. In addition, silencing of TIRAP effectively inhibited the activation of the Transforming Growth Factor-Beta-Activated Kinase 1/I-Kappa-B Kinase /Nuclear Factor of Kappa Light Polypeptide Gene Enhancer in B-Cells (TAK1/IKK/NF-κB) signalling pathway and the phosphorylation of Akt and mitogen-activated protein kinases, which were activated by LPS/IFN-γ stimulation. Thus, our results suggest that TIRAP positively regulated BV2 microglial M1 polarization through TLR4-mediated TAK1/IKK/NF-κB, mitogen-activated protein kinases and Akt signalling pathways.

Global analysis of transcriptome in dorsal root ganglia following peripheral nerve injury in rats.

Peripheral nervous system has intrinsic regeneration ability after injury, accompanied with the coordination of numerous cells, molecules and signaling pathways. These post-injury biological changes are complex with insufficient understanding. Thus, to obtain a global perspective of changes following nerve injury and to elucidate the mechanisms underlying nerve regeneration are of great importance. By RNA sequencing, we detected transcriptional changes in dorsal root ganglia (DRG) neurons at 0 h, 3 h, 9 h, 1 d, 4 d and 7 d following sciatic nerve crush injury in rats. Differentially expressed genes were then selected and classified into major clusters according to their expression patterns. Cluster 2 (with genes high expressed before 9 h and then down expressed) and cluster 6 (combination of cluster 4 and 5 with genes low expressed before 1 d and then up expressed) were underwent GO annotation and KEGG pathway analysis. Gene act networks were then constructed for these two clusters and the expression of pivotal genes was validated by quantitative real-time PCR. This study provided valuable information regarding the transcriptome changes in DRG neurons following nerve injury, identified potential genes that could be used for improving axon regeneration after nerve injury, and facilitated to elucidate the biological process and molecular mechanisms underlying peripheral nerve injury.

Identification and evaluation of the chemical similarity of Yindan xinnaotong samples by ultra high performance liquid chromatography with quadrupole time-of-flight mass spectrometry fingerprinting.

Yindan xinnaotong, a compound preparation used in traditional Chinese medicine, is composed of eight herbs: Ginkgo biloba leaf (yinxingye), Salvia miltiorrhizae (danshen), Herba gynostemmatis (jiaogulan), Erigerontis herba (dengzhanxixin), Allii sativi bulbus (dasuan), Notoginseng radixe rhizoma (sanqi), Crataegi fructus (shanzha), and Borneolum (tianranbingpian). Yindan xinnaotong is primarily used to treat cardiovascular and cerebrovascular diseases. However, to date, no scientific methods have been established to assess the quality of Yindan xinnaotong. Therefore, a combinatorial method was developed based on chemical constituent identification and fingerprint analysis to assess the consistency of Yindan xinnaotong quality. In this study, ultra high performance liquid chromatography coupled with time-of-flight mass spectrometry was used to identify the chemical components of Yindan xinnaotong soft capsules. Approximately 74 components were detected, of which 70, including flavonoids, ginkgolide, phenolic acid, diterpenoid tanshinones, and ginsenoside, were tentatively identified. A fingerprint analysis was also conducted to evaluate the uniformity of the quality of Yindan xinnaotong soft capsules. Ten batches of Yindan xinnaotong soft capsules were analyzed. All of the resulting chromatograms were imported into the "Similarity Evaluation System for Chromatographic Fingerprints of TCM" (Chinese Pharmacopoeia Commission, version 2004A). The similarity scores of common peaks from these samples ranged from 0.903-1.000, indicating that samples from different batches were highly correlated.

Prevention of atherosclerosis by Yindan Xinnaotong capsule combined with swimming in rats.

BACKGROUND: Yindan Xinnaotong capsule has been used for treating cardio-cerebrovascular diseases for several decades in China. Exercise training can protect against the development of atherosclerosis. The aim of the present study is to evaluate the joint effect of YXC and exercise on atherosclerosis in rats. METHODS: A combined method involving low shear stress and a high-fat diet was used to establish the atherosclerosis model in rats. Partial ligation of the left common carotid artery was performed, and then the rats were divided into 9 treatment groups according to a 3 × 3 factorial design with two factors and three levels for each factor, swimming of 0, 0.5, 1 h daily and YXC administration of 0, 1, 2 g/kg p.o. daily. Next the interventions of swimming and YXC were executed for 8 weeks. After that, blood samples were collected to determine blood viscosity, plasma viscosity, haematocrit (HCT), fibrinogen (FIB), blood lipid profile (including total cholesterol (TC), low-density lipoprotein-cholesterol (LDL-C), triglyceride (TG) and high-density lipoprotein-cholesterol (HDL-C)), nitric oxide (NO), 6-keto- prostaglandin (PG) F1α, endothelin (ET) and thromboxane (TX) B2. The common carotid arteries of the rats were harvested to examine pathological changes, wall thickness and circumference, and the expression of SM22αwas assayed via immune-histochemistry. RESULTS: The early pathological changes were observed. The joint effects of YXC and swimming showed significant changes in the examined parameters: (1) decreases in plasma viscosity, blood viscosity and FIB; (2) increases in NO and 6-keto-PGF1α; (3) decreases in ET and TXB2; and (4) decreases in LDL-C and TG. The combination of 2 g/kg YXC and 1 h of swimming led to synergistic decreases in LDL-C and TG. The interactive effect between YXC and swimming was obvious in decreasing wall thickness. Swimming alone was able to up-regulate the expression of SM22α. CONCLUSIONS: In conclusion, this study indicates that the combination of YXC and swimming may prevent atherosclerosis through a synergistic effect between YXC and swimming in improving blood circulation, hemorheological parameters, blood lipids levels and the vascular endothelium in rats. The vascular remodeling may be contributed to the prevention effects on AS by up-regulating SM22α.

Characterization and Analysis of Corrosion Resistance of Rubber Materials for Downhole Tools in a High-Stress Environment with Coupled H2S-CO2.

In the process of constructing deep natural gas wells in Sichuan and Chongqing, gas wells encounter various technical challenges such as high temperature, high pressure, and a corrosive environment containing H2S and CO2. The corrosion of rubber materials in these acidic environments can easily lead to seal failure in downhole tools. To better investigate the corrosion resistance of rubber materials in acidic environments, we utilized a dynamic cyclic corrosion experimental device capable of simulating the service conditions experienced by downhole tools under high-temperature, high-pressure multiphase flow. Corrosion-resistance tests were conducted on fluororubbers (FKM) 1, 2, 3, and HNBR (hydrogenated nitrile-butadiene rubber) under acidic conditions (80 °C and 160 °C), along with sealing corrosion tests on O-rings. These tests aimed to analyze the mechanical properties, hardness, and corrosion resistance before and after exposure to acid media as well as the sealing performance of O-rings. Ultimately, our goal was to identify suitable rubber materials for acidic pressure environments. Experimental results revealed that all four types of rubber exhibited decreased elongation at break after undergoing corrosion testing; however, fluororubber 3 demonstrated significant susceptibility to temperature effects while the other three types showed minimal impact from temperature variations. Fluororubber 1 and fluororubber 3 displayed substantial deformation levels whereas mechanical properties greatly deteriorated for fluororubber 2. Overall, HNBR showcased superior comprehensive performance.

Kuntai capsule attenuates premature ovarian insufficiency by activating the FOXO3/SIRT5 signaling pathway in mice: A comprehensive study using UHPLC-LTQ-Orbitrap and integrated pharmacology.

ETHNOPHARMACOLOGICAL RELEVANCE: Classical prescriptions are not only a primary method of clinical treatment in traditional Chinese medicine (TCM) but also represent breakthroughs in the inheritance and development of this field. Kuntai capsule (KTC), a formulation based on a classical prescription, comprises six TCMs: Rehmanniae Radix Praeparata, Coptidis Rhizoma, Paeoniae Radix Alba, Scutellariae Radix, Asini Corii Colla, and Poria. This formulation possesses various beneficial effects, such as nourishing yin and blood, clearing heat and purging fire, and calming the nerves and relieving annoyance. The investigation of the efficacy and mechanism of KTC in regulating anti-aging factors in the treatment of premature ovarian insufficiency (POI) is not only a prominent topic in classical prescription research but also a crucial issue in the treatment of female reproductive aging using TCM. AIM OF THE STUDY: To evaluate the therapeutic effect of KTC on POI and its underlying mechanism. MATERIALS AND METHODS: Healthy and specific pathogen-free (SPF) female Kunming mice aged 6-8 weeks were selected. After acclimatization, the mice were randomly divided into a control, model, and high, middle, and low dose groups of KTC (1.6, 0.8, and 0.4 mg/kg, respectively). Except for the control group, the animals in the other groups were administered a single intraperitoneal injection of 120 mg/kg cyclophosphamide and 30 mg/kg Busulfan to induce the model of POI. After modeling, the mice were treated with the corresponding drugs for 7 days. Serum and ovarian tissues were collected, and the levels of serum follicle-stimulating hormone (FSH), estradiol (E2), and superoxide dismutase 2 (SOD2) were determined using enzyme-linked immunosorbent assay (ELISA). The chemical composition of KTC was characterized and analyzed using ultra-high-pressure liquid chromatography-linear ion trap-Orbitrap tandem mass spectrometry. A "drug-component-target-pathway-disease" network was constructed using network pharmacology research methods to identify the key active components of KTC in treating POI and to elucidate its potential mechanism. The protein expression of the FOXO3/SIRT5 pathway was detected by western blotting. RESULTS: Compared to the model group, the high-dose group of KTC showed a significant increase in ovarian index, significant increase in levels of E2 and SOD2, and a significant decrease in FSH levels. Through systematic analysis of the chemical constituents of KTC, 69 compounds were identified, including 7 organic acids, 14 alkaloids, 28 flavonoids, 15 terpenoids, 2 lignans, 2 phenylpropanoids, and 1 sugar. Based on network pharmacology research methods, it was determined that KTC exerts its therapeutic effect on POI through multiple components (paeoniflorin and malic acid), multiple targets (FOXO3 and SIRT5), and multiple pathways (prolactin signaling pathway, longevity regulating pathway, and metabolic pathways). The accuracy of the network pharmacology prediction was further validated by detecting the protein expression of SIRT5 and FOXO3a, which showed a significant increase in the middle and high-dose groups of KTC compared to the model group. CONCLUSIONS: KTC may effectively treat POI through a multi-component, multi-target, multi-pathway approach, providing an experimental basis for using KTC based on classical prescriptions in the treatment of POI.

Intranasal Administration of Umbilical Cord Mesenchymal Stem Cell Exosomes Alleviates Parkinson's Disease.

Parkinson's disease (PD) is a common and complex neurodegenerative disease. This disease is typically characterized by the formation of Lewy bodies in multiple brain regions and dopaminergic neuronal loss in the substantia nigra pars compacta, resulting in non-motor symptoms (e.g., olfactory deficits) and motor dysfunction in the late stages. There is yet no effective cure for Parkinson's disease. Considering the neuroprotective effects of exosomes, we investigated whether intranasal administration of umbilical cord mesenchymal stem cell exosomes could improve behavioral functions in PD mice. First, exosomes were endocytosed by the cells in vitro and in vivo, indicating that exosomes can cross the blood-brain barrier. Second, we found that both motor and non-motor functions of the PD models were effectively improved during intranasal exosomes treatment. Finally, the activity of olfactory bulb neurons was improved and the loss of dopaminergic neurons in the substantia nigra pars compacta was reversed. Moreover, exosomes attenuated microglia and astrocyte activation, leading to a low level of inflammation in the brain. In conclusion, our study provided a new reference for the clinical application of exosomes in the treatment of PD.

Single-Cell RNA Sequencing Analysis of Microglia Dissected the Energy Metabolism and Revealed Potential Biomarkers in Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) is a common neurodegenerative disease, accompanied by the gradual loss of motor neuron, even life-threatening. However, the pathogenesis, early diagnosis, and effective strategies of ALS are not yet completely understood. In this study, the function of differentially expressed genes (DEGs) in non-neuronal cells of the primary motor cortex of ALS patients (DATA1), the brainstem of SOD1 mutant ALS mice (DATA2), and the whole blood tissue of ALS patients (DATA3) were explored. The results showed that the functions of DEGs in non-neuronal cells were mainly related to energy metabolism (such as oxidative phosphorylation) and protein synthesis. In non-neuronal cells, six upregulated DEGs (HSPA8, SOD1, CALM1, CALM2, NEFL, COX6C) and three downregulated DEGs (SNRNP70, HSPA1A, HSPA1B) might be key factors in regulating ALS. Microglia played a key role in the development of ALS. The expression of SOD1 and TUBA4A in microglia in DATA1 was significantly increased. The integration analysis of DEGs in DATA1 and DATA2 showed that SOD1 and CALM1 might be potential biomarkers. The integration analysis of DEGs in DATA1 and DATA3 showed that CALM2 and HSPA1A might be potential biomarkers. Cell interaction showed that the interaction between microglia and other cells was reduced in high oxidative phosphorylation states, which might be a risk factor in ALS. Our research provided evidence for the pathogenesis, early diagnosis, and potential targeted therapy for ALS.

Skin derived precursors induced Schwann cells mediated tissue engineering-aided neuroregeneration across sciatic nerve defect.

A central question in neural tissue engineering is how the tissue-engineered nerve (TEN) translates detailed transcriptional signals associated with peripheral nerve regeneration into meaningful biological processes. Here, we report a skin-derived precursor-induced Schwann cell (SKP-SC)-mediated chitosan/silk fibroin-fabricated tissue-engineered nerve graft (SKP-SCs-TEN) that can promote sciatic nerve regeneration and functional restoration nearly to the levels achieved by autologous nerve grafts according to behavioral, histological, and electrophysiological evidence. For achieving better effect of neuroregeneration, this is the first time to jointly apply a dynamic perfusion bioreactor and the ascorbic acid to stimulate the SKP-SCs secretion of extracellular matrix (ECM). To overcome the limitation of traditional tissue-engineered nerve grafts, jointly utilizing SKP-SCs and their ECM components were motivated by the thought of prolongating the effect of support cells and their bioactive cues that promote peripheral nerve regeneration. To further explore the regulatory model of gene expression and the related molecular mechanisms involved in tissue engineering-aided peripheral nerve regeneration, we performed a cDNA microarray analysis of gene expression profiling, a comprehensive bioinformatics analysis and a validation study on the grafted segments and dorsal root ganglia tissues. A wealth of transcriptomic and bioinformatics data has revealed complex molecular networks and orchestrated functional regulation that may be responsible for the effects of SKP-SCs-TEN on promoting peripheral nerve regeneration. Our work provides new insights into transcriptomic features and patterns of molecular regulation in nerve functional recovery aided by SKP-SCs-TEN that sheds light on the broader possibilities for novel repair strategies of peripheral nerve injury.

MircoRNA-25-3p in skin precursor cell-induced Schwann cell-derived extracellular vesicles promotes axon regeneration by targeting Tgif1.

Nerve injury often leads to severe dysfunction because of the lack of axon regeneration in adult mammal. Intriguingly a series of extracellular vesicles (EVs) have the obvious ability to accelerate the nerve repair. However, the detailed molecular mechanisms to describe that EVs switch neuron from a transmitter to a regenerative state have not been elucidated. This study elucidated the microRNA (miRNA) expression profiles of two types of EVs that promote nerve regeneration. The functions of these miRNAs were screened in vitro. Among the 12 overlapping miRNAs, miR-25-3p was selected for further analysis as it markedly promoted axon regeneration both in vivo and in vitro. Furthermore, knockdown experiments confirmed that PTEN and Klf4, which are the major inhibitors of axon regeneration, were the direct targets of miR-25-3p in dorsal root ganglion (DRG) neurons. The utilization of luciferase reporter assays and functional tests provided evidence that miR-25-3p enhances axon regeneration by targeting Tgif1. Additionally, miR-25-3p upregulated the phosphorylation of Erk. Furthermore, Rapamycin modulated the expression of miR-25-3p in DRG neurons. Finally, the pro-axon regeneration effects of EVs were confirmed by overexpressing miR-25-3p and Tgif1 knockdown in the optic nerve crush model. Thus, the enrichment of miR-25-3p in EVs suggests that it regulates axon regeneration, proving a potential cell-free treatment strategy for nerve injury.

Combined gas well hydrate prevention and control technology and its application.

The high pressure in some gas wells, such as those in the Xushen gas field in Daqing, China, makes them susceptible to freezing and hydrate blockages. Downhole throttling technology is widely used to reduce costs during well construction, however, due to the limitations of temperature, pressure and depth structure, this technology is sometime applied independently in some gas wells in which freezing and blockages are a frequent problem that can seriously affect production capacity. Moreover, artificial alcohol injection of 'passive plugging' to prevent hydrate formation not only consumes significant amounts of methanol but its efficiency is also dependent on factors such as weather, personnel and equipment, so it is not a continuous solution. In order to solve the above problems, the mechanism of hydrate formation was analyzed in this study, from which a combined mechanical and chemical hydrate control process was developed. OLGA software was used to design the process parameters of the novel mechanical and chemical inhibition technology for hydrate prevention and control, and also to simulate and analyze the wellhead temperature, pressure and hydrate generation once the process was implemented. Based on the results of the parameters calculation, the downhole throttle and hydrate inhibitor automatic filling device are used to realize the functions of downhole throttle depressurization and hydrate inhibitor continuous filling, reduce the wellhead pressure and hydrate generation temperature, and ensure the continuous production of gas well. This novel combination process was subsequently tested in three wells in the Daqing gas oilfield. Measurements showed that the average daily gas increase from a single well was 0.5×104m3, methanol consumption was reduced from the original maximum daily amount of 1750 kg to just 60 kg, the manual maintenance workload was reduced by 80%, and the rate of the well openings was increased from 45% to 100%. These results proved that this technology is feasible and efficient for applications in gas wells with high downhole pressure and low wellhead temperature, and, thus, provides important technical support for the prevention of gas hydrate and improvement of gas well production.