ObjectiveTo observe the changes in the levels of different subtypes of epidermal growth factor receptor （ErbB）， namely ErbB1， ErbB2， ErbB3， and ErbB4， in the spinal dorsal horn of inflammatory pain model rats， and to explore their mechanism of mediating hyperalgesia as well as the intervention mechanism of electroacupuncture at "Zusanli （ST 36）" and "Kunlun （BL 60）". MethodsThe study was divided into five parts. In experiment 1， 14 Sprague Dawley （SD） rats were randomly divided into control and inflammatory pain group （7 rats each group） to observe the pain behavior and the protein expression of different ErbB receptor subtypes in the spinal dorsal horn. In experiment 2， 30 rats were randomly divided into control group 1， inflammatory pain group 1， and low-， medium-， and high-concentration TX1-85-1 groups， with 6 rats in each group， to observe the effect of inhibiting spinal ErbB3 on inflammatory pain. In experiment 3， 12 rats were randomly divided into control virus group and ErbB3 knockdown virus group， with 6 rats in each group， to observe the effect of knocking down ErbB3 in the spinal dorsal horn on inflammatory pain. In experiment 4， 44 rats were randomly divided into control group 2， inflammatory pain group 2， electroacupuncture group， and sham electroacupuncture group， with 11 rats in each group， to observe the effect of electroacupuncture. In experiment 5， 40 rats were randomly divided into control group 3， inflammatory pain group 3， electroacupuncture group 1， and electroacupuncture + NRG1 group， with 10 rats in each group， to observe the effect of activating ErbB3 on electroacupuncture. A rat model of inflammatory pain was established by subcutaneous injection of 100 μl of complete Freund's adjuvant into the sole of the unilateral hind foot of SD rats. Rats in the low-， medium-， and high-concentration TX1-85-1 groups were intrathecally injected with ErbB3 inhibitor TX1-85-1 on day 5 to day 7 after modeling. Rats in the ErbB3 knockdown virus group were injected with ErbB3 knockdown virus packaged with adenovirus vector-based short hairpin RNA （shRNA） into the spinal dorsal horn in situ 3 weeks before modeling. Rats in each electroacupuncture group received electroacupuncture at bilateral "Zusanli （ST 36）" and "Kunlun （BL 60）" from day 1 to day 7 after modeling， with dense-sparse waves at a frequency of 2 Hz/100 Hz and a current of 0.5-1.5 mA for 30 minutes once a day. Rats in the electroacupuncture + NRG1 group were intrathecally injected with ErbB3 ligand recombinant human neuregulin-1 （NRG1） after electroacupuncture intervention from day 5 to day 7 after modeling. The mechanical withdrawal threshold and thermal withdrawal latency of rats were measured on day 1， 3， 5， and 7 after modeling to evaluate behavior， and Western Blot was used to detect the protein and phosphorylation levels of each ErbB subtype in the spinal dorsal horn. ResultsCompared with the control group， rats in the inflammatory pain group showed decreased mechanical withdrawal threshold and thermal withdrawal latency of rats， and increased expression of phosphorylated ErbB3 （p-ErbB3） protein in the spinal dorsal horn on days 1， 3， 5， and 7 after modeling （P＜0.01）. On day 5 and day 7 after modeling， compared with the inflammatory pain group 1， the mecha-nical withdrawal threshold and thermal withdrawal latency of rats in the medium- and high-concentration TX1-85-1 groups increased， and the expression of p-ErbB3 protein decreased （P＜0.05）. On day 1， 3， 5， and 7 after modeling， compared with the control virus group， the mechanical withdrawal threshold and thermal withdrawal latency of rats in the ErbB3 knockdown virus group increased （P＜0.05）. On day 5 and day 7 after modeling， compared with the inflammatory pain group 2 and the sham electroacupuncture group， the mechanical withdrawal threshold and thermal withdrawal latency of rats in the electroacupuncture group increased， and the expression of p-ErbB3 protein decreased （P＜0.05）. On day 5 and day 7 after modeling， compared with the electroacupuncture + NRG1 group， the mechanical withdrawal threshold and thermal withdrawal latency of rats in the electroacupuncture group 1 increased （P＜0.05）. ConclusionThe p-ErbB3 in the spinal dorsal horn involved in hyperalgesia in rats with inflammatory pain， and electroacupuncture at "Zusanli （ST 36）" and "Kunlun （BL 60）" can alleviate inflammatory pain by inhibiting the expression of p-ErbB3 protein in the spinal dorsal horn of rats.

ObjectiveTo investigate the gene expression sequence and molecular mechanisms in the local microenvironment during the subacute to chronic phases (1-28 days) in mouse models of spinal cord compression injury and hemisection spinal cord injury, thereby revealing the molecular characteristics of spinal cord repair and providing a theoretical basis for selecting therapeutic targets for spinal cord injury. MethodsThirty-six 8-9-week-old SPF-grade ICR mice were randomly divided into three groups (n=12 per group): sham-operated control (CTR) group, hemisection spinal cord injury (HSCI) group, and spinal cord compression injury (SCC) group. Mice in the CTR group underwent the same surgical preparation and anesthesia, followed by a dorsal midline incision at the T₉-T₁₀ segment. After layer-by-layer dissection and removal of the corresponding lamina, the spinal cord dura mater was fully exposed and kept intact. The cord was exposed to air for 10 minutes (matching the duration of the compression injury group), during which any instrument contact with the cord was avoided. The incision was then irrigated and sutured. The HSCI group underwent a 70% transection of the T₉ spinal cord segment using micro-instruments to establish a hemisection spinal cord injury model. The SCC group underwent sustained compression of the T₁₀ spinal cord segment for 10 minutes using a self-made compressor (a 30 g solid small iron bar) to establish a spinal cord compression injury model. Motor function recovery was assessed using the modified Basso-Beattie-Bresnahan (BBB) score on postoperative days 1, 3, 7, 14, 21, and 28. On days 7 and 14 post-operation, mice were anesthetized, and the injured spinal cord segments were harvested. The evolution of specific molecular networks in the spinal cord injury mouse models was analyzed via RNA sequencing (RNA-Seq) and enrichment analysis, and the expression of key genes was verified using real time fluorogenic quantitative PCR. ResultsBBB scores indicated that motor function recovery in the SCC group was significantly better than that in the HSCI group, with BBB scores showing a continuously increasing trend and remaining higher than those in the HSCI group over the 4-week period (P <0.001). Gene ontology （GO)and Kyoto Encyclopedia of Genes and Genomes （KEGG） enrichment analyses based on RNA-Seq differentially expressed genes revealed that, compared to the CTR group, genes related to the extracellular matrix were significantly up-regulated (P<0.05), while genes related to axon guidance were significantly down-regulated (P <0.05) in the SCC group on day 7 post-operation. On day 21, genes involved in immune regulation and the retinol signaling pathway were significantly activated in the SCC group (P<0.05). In contrast, in the HSCI group, genes associated with inflammation and immune response were significantly up-regulated (P<0.001), while genes related to neuronal differentiation and synapse formation were significantly down-regulated (P <0.001) on day 7. On day 21, genes related to cell-matrix junctions and N-methyl-D-aspartate receptors were significantly up-regulated (P<0.001) in the HSCI group. Furthermore, compared to the SCC group, the HSCI group exhibited different pathway enrichment characteristics in GO and KEGG analyses on days 7 and 21 post-injury. On day 7, genes involved in the NOD-like receptor signaling pathway and the complement and coagulation cascades were significantly up-regulated in the HSCI group (P<0.001). On day 21, genes related to the extracellular matrix-receptor interaction and the neuroactive ligand-receptor interaction pathways were significantly activated (P<0.001). Finally, real time fluorogenic quantitative PCR validation results were highly consistent with the RNA-Seq results, further confirming the differential expression trends of key genes between the SCC and HSCI groups. ConclusionThe SCC and HSCI injury models may drive distinct repair pathways: the preservation of some axons in the SCC model predisposes it toward tissue repair, whereas the HSCI model requires the coordination of more complex molecular networks to achieve a new equilibrium. This finding further deepens the understanding of the heterogeneous regulatory mechanisms underlying spinal cord injury.

Objective: To elucidate the expression level and transcript structure of the CD36 gene in megakaryocytes cultured in vitro. Methods: Using umbilical cord blood CD34 hematopoietic stem cells as the starting point, megakaryocytic lineage cells were directionally cultured in vitro using different combinations of cytokines. Total cell RNA was extracted from cultures at 0 d, 7 d, and 12 d, and megakaryocyte RNA was extracted from CD41a-sorted cells after 14 d and 18 d of culture. RNA-NGS sequencing technology was used to analyze the RNA gene expression profiles across the five culture periods and further investigate CD36 gene expression. Results: The number of megakaryocytes generated in the TPO (100 ng/mL) group [(2.2±0.02)×10 /mL] was significantly higher than that in the other five groups. A total of 22 066 expressed genes were detected in the RNA of the five culture periods, and gene expression levels at each time point were correlated with the culture timeline. CD36 gene expression increased with culture time, with FPKM values for CD36 expression in the megakaryocytic lineage at 14 d and 18 d being 18.35 and 101.85, respectively, which were much lower than those for ITGA2B and ITGB3 genes but slightly higher than that for CD109 gene in the megakaryocytic lineage. The longest transcript of CD36 in the 18 d megakaryocytic lineage was 3.8 kb, encompassing all sequence of exons E3 to E14 and a partial sequence of E15. Conclusion: This is the first report on the expression level and transcript structure of the CD36 gene in megakaryocytes cultured in vitro, providing fundamental data for research on the expression and regulation of the CD36 gene in the megakaryocytic lineage.

Background/Aims: Metabolic dysfunction–associated steatotic liver disease (MASLD) is a chronic liver disease characterized by hepatic steatosis. Ubiquitin-specific protease 29 (USP29) plays pivotal roles in hepatic ischemiareperfusion injury and hepatocellular carcinoma, but its role in MASLD remains unexplored. Therefore, the aim of this study was to reveal the effects and underlying mechanisms of USP29 in MASLD progression. Methods: USP29 expression was assessed in liver samples from MASLD patients and mice. The role and molecular mechanism of USP29 in MASLD were assessed in high-fat diet-fed and high-fat/high-cholesterol diet-fed mice and palmitic acid and oleic acid treated hepatocytes. Results: USP29 protein levels were significantly reduced in mice and humans with MASLD. Hepatic steatosis, inflammation and fibrosis were significantly exacerbated by USP29 deletion and relieved by USP29 overexpression. Mechanistically, USP29 significantly activated the expression of genes related to fatty acid β-oxidation (FAO) under metabolic stimulation, directly interacted with long-chain acyl-CoA synthase 5 (ACSL5) and repressed ACSL5 degradation by increasing ACSL5 K48-linked deubiquitination. Moreover, the effect of USP29 on hepatocyte lipid accumulation and MASLD was dependent on ACSL5. Conclusions USP29 functions as a novel negative regulator of MASLD by stabilizing ACSL5 to promote FAO. The activation of the USP29-ACSL5 axis may represent a potential therapeutic strategy for MASLD.

Purpose: To investigate the therapeutic potential of eliminating insulin-like growth factor-binding protein 5 (IGFBP5) expression in improving erectile function in mice with cavernous nerve injury (CNI)-induced erectile dysfunction (ED). Materials and Methods: Eight-week-old male C57BL/6 mice were divided into four groups: a sham-operated group and three CNI-induced ED groups. The CNI-induced ED groups were treated with intracavernous injections 3 days before the CNI procedure.These injections included phosphate-buffered saline, scrambled control short hairpin RNA (shRNA), or shRNA targeting mouse IGFBP5 lentiviral particles. One week after CNI, erectile function was evaluated and the penile tissue was then harvested for histological examination and western blot analysis. Additionally, the major pelvic ganglia (MPG) and dorsal root ganglia (DRG) were cultured for ex vivo neurite outgrowth assays. Results: Following CNI, IGFBP5 expression in the cavernous tissues significantly increased, reaching its peak at day 7. First, ablation of IGFBP5 expression promotes neurite sprouting in MPG and DRG when exposed to lipopolysaccharide. Second, ablating IGFBP5 expression in CNI-induced ED mice improved erectile function, likely owing to increased neurovascular contents, including endothelial cells, pericytes, and neuronal processes. Third, ablating IGFBP5 expression in CNI-induced ED mice promoted neurovascular regeneration by increasing cell proliferation, reducing apoptosis, and decreasing Reactive oxygen species production. Finally, western blot analysis demonstrated that IGFBP5 ablation attenuated the JNK/c-Jun signaling pathway, activated the PI3K/AKT signaling pathway, and increased vascular endothelial growth factor and neurotrophic factor expression. Conclusions Ablating IGFBP5 expression enhanced neurovascular regeneration and ultimately improved erectile function in CNI-induced ED mice.

Background/Aims: Metabolic dysfunction–associated steatotic liver disease (MASLD) is a chronic liver disease characterized by hepatic steatosis. Ubiquitin-specific protease 29 (USP29) plays pivotal roles in hepatic ischemiareperfusion injury and hepatocellular carcinoma, but its role in MASLD remains unexplored. Therefore, the aim of this study was to reveal the effects and underlying mechanisms of USP29 in MASLD progression. Methods: USP29 expression was assessed in liver samples from MASLD patients and mice. The role and molecular mechanism of USP29 in MASLD were assessed in high-fat diet-fed and high-fat/high-cholesterol diet-fed mice and palmitic acid and oleic acid treated hepatocytes. Results: USP29 protein levels were significantly reduced in mice and humans with MASLD. Hepatic steatosis, inflammation and fibrosis were significantly exacerbated by USP29 deletion and relieved by USP29 overexpression. Mechanistically, USP29 significantly activated the expression of genes related to fatty acid β-oxidation (FAO) under metabolic stimulation, directly interacted with long-chain acyl-CoA synthase 5 (ACSL5) and repressed ACSL5 degradation by increasing ACSL5 K48-linked deubiquitination. Moreover, the effect of USP29 on hepatocyte lipid accumulation and MASLD was dependent on ACSL5. Conclusions USP29 functions as a novel negative regulator of MASLD by stabilizing ACSL5 to promote FAO. The activation of the USP29-ACSL5 axis may represent a potential therapeutic strategy for MASLD.

Background Metals and metalloids in fine particulate matter (PM_2.5) may cause damage to the respiratory and circulatory systems of the human body, and long-term exposure is prone to causing chronic poisoning, cancer, and other adverse effects. Objective To assess the distribution characteristics of metals and metalloids in outdoor PM_2.5 in a southern city of China, conduct source apportionment, and evaluate the associated health risks, thereby providing theoretical support for further pollution control measures. Methods PM_2.5 samples were collected in districts A, B, and C of a southern China city, and the concentrations of 17 metals and metalloids were detected by inductively coupled plasma-mass spectrometry (ICP-MS). Pollution sources were assessed through enrichment factor and principal components analysis, and the main pollution sources were quantified using absolute principal component scores-multivariate linear regression (APCS-MLR). Health risks were evaluated based on the Technical guide for environmental health risk assessment of chemical exposure (WS/T777—2021). Results The ambient air PM_2.5 concentrations in the city were higher in winter and spring, and lower in summer and autumn. The annual average concentrations of ambient PM_2.5 in districts A, B, and C were 36.7, 31.9, and 24.4 μg·m⁻³, respectively. The ambient PM_2.5 levels in districts B and C were below the second-grade limit set by the Ambient air quality standards (GB 3095—2012). The enrichment factors of cadmium (Cd), aluminum (Al), and antimony (Sb) were greater than 10, those of copper (Cu), lead (Pb), arsenic (As), nickel (Ni), mercury (Hg), and molybdenum (Mo) fell between 1 and 10, and those of manganese (Mn), vanadium (V), chromium (Cr), cobalt (Co), barium (Ba), beryllium (Be), and uranium (U) were below or equal to 1. The comprehensive evaluation of source analysis showed that the main pollution sources in districts A and C and the whole city were coal-burning. In district B, the main pollution source was also coal combustion, followed by industrial process sources and dust sources. The carcinogenic risks of As and Cr were between 1×10⁻⁶ and 1×10⁻⁴. However, the hazard quotients for 15 metals and metalloids in terms of non-carcinogenic risk were below 1. Conclusion Cr and As in the atmospheric PM_2.5 of the city present a certain risk of cancer and should be paid attention to. In addition, preventive control measures should be taken against relevant pollution sources such as industrial emission, dust, and coal burning.

Background/Aims: Metabolic dysfunction–associated steatotic liver disease (MASLD) is a chronic liver disease characterized by hepatic steatosis. Ubiquitin-specific protease 29 (USP29) plays pivotal roles in hepatic ischemiareperfusion injury and hepatocellular carcinoma, but its role in MASLD remains unexplored. Therefore, the aim of this study was to reveal the effects and underlying mechanisms of USP29 in MASLD progression. Methods: USP29 expression was assessed in liver samples from MASLD patients and mice. The role and molecular mechanism of USP29 in MASLD were assessed in high-fat diet-fed and high-fat/high-cholesterol diet-fed mice and palmitic acid and oleic acid treated hepatocytes. Results: USP29 protein levels were significantly reduced in mice and humans with MASLD. Hepatic steatosis, inflammation and fibrosis were significantly exacerbated by USP29 deletion and relieved by USP29 overexpression. Mechanistically, USP29 significantly activated the expression of genes related to fatty acid β-oxidation (FAO) under metabolic stimulation, directly interacted with long-chain acyl-CoA synthase 5 (ACSL5) and repressed ACSL5 degradation by increasing ACSL5 K48-linked deubiquitination. Moreover, the effect of USP29 on hepatocyte lipid accumulation and MASLD was dependent on ACSL5. Conclusions USP29 functions as a novel negative regulator of MASLD by stabilizing ACSL5 to promote FAO. The activation of the USP29-ACSL5 axis may represent a potential therapeutic strategy for MASLD.

Compounds isolated from Epimedium include the total flavonoids of Epimedium , icariin, and its metabolites (icaritin, icariside I, and icariside II), which have similar molecular structures. Modern pharmacological research and clinical practice have proved that Epimedium and its active components have a wide range of pharmacological effects, especially in improving sexual function, hormone regulation, anti-osteoporosis, immune function regulation, anti-oxidation, and anti-tumor activity. To date, we still need a comprehensive source of knowledge about the pharmacological effects of Epimedium and its bioactive compounds on the male reproductive system. However, their actions in other tissues have been reviewed in recent years. This review critically focuses on the Epimedium , its bioactive compounds, and the biochemical and molecular mechanisms that modulate vital pathways associated with the male reproductive system. Such intrinsic knowledge will significantly further studies on the Epimedium and its bioactive compounds that protect the male reproductive system and provide some guidances for clinical treatment of related male reproductive disorders.
Male ; Epimedium/chemistry* ; Humans ; Genitalia, Male/drug effects* ; Flavonoids/therapeutic use* ; Animals

Prostate cancer is the second most common malignancy and the sixth leading cause of cancer-related death in men worldwide. Radical prostatectomy (RP) is the standard treatment for localized prostate cancer, but the procedure often results in postoperative erectile dysfunction (ED). The poor efficacy of phosphodiesterase 5 inhibitors after surgery highlights the need to develop new therapies to enhance cavernous nerve regeneration and improve the erectile function of these patients. In the present study, we aimed to examine the potential of heparin-binding epidermal growth factor-like growth factor (HB-EGF) in preserving erectile function in cavernous nerve injury (CNI) mice. We found that HB-EGF expression was reduced significantly on the 1 st day after CNI in penile tissue. Ex vivo and in vitro studies showed that HB-EGF promotes major pelvic ganglion neurite sprouting and neuro-2a (N2a) cell migration. In vivo studies showed that exogenous HB-EGF treatment significantly restored the erectile function of CNI mice to 86.9% of sham levels. Immunofluorescence staining showed that mural and neuronal cells were preserved by inducing cell proliferation and reducing apoptosis and reactive oxygen species production. Western blot analysis showed that HB-EGF upregulated protein kinase B and extracellular signal-regulated kinase activation and neurotrophic factor expression. Overall, HB-EGF is a major promising therapeutic agent for treating ED in postoperative RP.
Animals ; Male ; Heparin-binding EGF-like Growth Factor/therapeutic use* ; Erectile Dysfunction/etiology* ; Mice ; Penis/drug effects* ; Nerve Regeneration/drug effects* ; Penile Erection/drug effects* ; Peripheral Nerve Injuries/drug therapy* ; Cell Proliferation/drug effects* ; Apoptosis/drug effects* ; Cell Movement/drug effects* ; Prostatectomy/adverse effects* ; Mice, Inbred C57BL ; Reactive Oxygen Species/metabolism*