Background: Influenza A virus (IAV) is a negative-sense RNA virus of the Orthomyxoviridae family and is the etiological agent of a highly contagious acute respiratory disease that can lead to acute lung injury. Objective: To elucidate the molecular mechanisms of IAV infection, an integrative research approach combining gene expression profiling, multinetwork analysis, and in vivo experimental validations was employed. Methods: First, a series of network-based analyses were performed, including protein-protein interaction network construction, weighted gene co-expression network analysis, and subsequent gene set enrichment analysis, to identify the major underlying mechanisms of IAV infection. Following gene expression analysis, core targets, both direct and indirect regulators, were screened. An IAV (H1N1) strain A/PR/8/34-induced acute lung injury mouse model was constructed for in vivo validations. Batch one included two groups to evaluate findings from the multi-network analysis: Mock (n = 10; 5 males and 5 females) and IAV (n = 10; 5 males and 5 females). Batch two included three groups to assess the role of toll-like receptor 3 (TLR3) in IAV infection: Mock (n = 6; 3 males and 3 females), IAV (n = 6; 3 males and 3 females), and TLR3 inhibitor (n = 6; 3 males and 3 females). Body weight was measured on days 0, 3, and 5 after infection. On day 5, lung tissues were collected to assess viral load and histopathological changes. Key targets were examined using enzyme-linked immunosorbent assay, Western blotting, and immunofluorescence staining, both in sera and lung tissues. Results: IAV infection was significantly associated with dysregulation of the immune-inflammation system, such as the LTR, nucle-otide-binding oligomerization domain-(NOD) like receptor, retinoic acid-inducible gene I-like receptor, and nuclear factor kappa-B signaling pathways. Gene set enrichment analysis further indicated that the TLR and necroptosis signaling pathways played crucial roles in the progression of IAV infection (TLR signaling pathway normalized enrichment score = 2.3941, P = 1.00 × 10 ⁻¹⁰; necroptosis normalized enrichment score = 1.9421, P = 6.21 × 10 ⁻⁷). Among the core targets, TLR3 and mixed lineage kinase domain-like protein (MLKL) may regulate gene expression at the transcriptional level (all P < 0.05). In vivo validation using an IAV (PR8) infected acute lung injury mouse model demonstrated increased viral load and lung index, alveolar structural damage, and inflammatory cell infiltration. Immunofluorescence staining exhibited large gaps in Lamin B1 staining and breaches in Emerin signals following IAV-PR8 infection. Expression levels of TLR3, p-receptor-interacting serine/threonine-protein kinase 3 (RIPK3)/RIPK3, and p-mixed lineage kinase domain-like protein (MLKL)/MLKL proteins in lung tissues, as well as proinflammatory factors and mediators in sera, were significantly elevated after IAV infection. Moreover, enhanced neutrophil infiltration (myeloperoxidase) and citrullinated histone H3 (a neutrophil extracellular trap-specific marker), both established indicators of neutrophil extracellular trap formation, were observed. Notably, treatment with a TLR3 inhibitor significantly ameliorated IAV-induced acute lung injury by regulating necroptosis-related targets. Conclusion: Our study provides network-based in vivo evidence that TLR3-receptor-interacting serine/threonine-protein kinase 3-MLKL-mediated necroptosis may underlie IAV-induced acute lung injury and could serve as a potential therapeutic target in severe influenza cases.

Objective To investigate the post-discharge exercise behavior and factors influencing moderate to vigorous intensity physical activity (MVPA) in patients undergoing lung surgery. Methods A total of 2874 patients from the large prospective, observational perioperative lung symptom study cohort (CN-PRO-Lung 3) in the Department of Thoracic Surgery at Sichuan Cancer Hospital between April 7, 2021, and January 31, 2024, were selected as the survey subjects. A survey was conducted using the Investigation of Exercise Behavior after Lung Surgery questionnaire and the International Physical Activity Questionnaire-Short Form (IPAQ-SF) among patients who underwent lung surgery. Binary logistic regression was used to analyze the factors influencing patients’ engagement in MVPA. Results A total of 702 patients were surveyed, including 252 males and 450 females, with an average age of (52.4±10.2) years. Patients with lung cancer accounted for 85.9%. Only 36.0% of the patients had regular exercise habits, while 42.3% did not engage in any physical activity. The three main barriers for postoperative exercise were physical discomfort (pain, coughing, shortness of breath, etc, 54.7%), lack of professional guidance (41.7%), and concerns about the surgical wound (28.9%). The proportions of patients engaging in vigorous, moderate, and low-intensity physical activity were 5.7%, 28.2%, and 66.1%, respectively. Multivariate analysis showed that patients with a personal annual income ≥50000 yuan (OR=1.52, 95%CI 1.01-2.29, P=0.044), high school education or above (OR=1.92, 95%CI 1.33-2.76, P<0.001), and lobectomy (OR=1.44, 95%CI 1.02-2.03, P=0.037) engaged in more MVPA. Conclusion Patients undergoing lung surgery have inadequate physical activity after discharge, particularly lacking in MVPA. Patients with higher income, higher educational levels, and lobectomy are more frequently engaged in MVPA. Measures such as symptom control, providing exercise guidance, and enhancing education on wound care may potentially improve the inadequate physical activity in lung surgery patients after discharge.

ObjectiveTo elucidate the underlying mechanism through which Zhuluan decoction suppresses excessive autophagy in human ovarian granulosa cells （KGN） and ameliorates premature ovarian insufficiency （POI） via the phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/mammalian target of rapamycin （mTOR） signaling pathway. MethodsThe optimal concentration of cyclophosphamide for inducing a POI model in KGN cells was identified via the cell counting kit-8 （CCK-8） assay. Subsequently， the impacts of varying concentrations of Zhuluan decoction-containing serum on the viability of the KGN cell model were assessed. After the optimal drug concentration was determined， KGN cells were categorized into the following groups： blank control （20% blank serum）， model （20% blank serum + 5 μmol·L^-1 cyclophosphamide）， Zhuluan decoction-containing serum （20% Zhuluan decoction-containing serum + 5 μmol·L^-1 cyclophosphamide）， autophagy inhibitor （20% blank serum + 5 μmol·L^-1 cyclophosphamide + 20 μmol·L^-1 chloroquine phosphate）， autophagy inhibitor + Zhuluan decoction-containing serum （20% Zhuluan decoction-containing serum + 5 μmol·L^-1 cyclophosphamide + 20 μmol·L^-1 chloroquine phosphate）， and estradiol valerate （20% estradiol valerate-containing serum + 5 μmol·L^-1 cyclophosphamide）. Following 48 hours of incubation， flow cytometry was utilized to measure the apoptosis rate of KGN cells in each group. Western blotting was employed to quantify the protein levels of PI3K， phosphorylated （p）-Akt， Akt， p-mTOR， and mTOR， along with the expression levels of autophagy-related proteins such as Beclin1， autophagy-related 5 homolog （ATG5）， and microtubule-associated protein 1 light chain 3 （LC3）， in each group. Additionally， monodansylcadaverine （MDC） staining was performed to evaluate the extent of autophagy in each group. ResultsIncubation of KGN cells with 5 μmol·L^-1 cyclophosphamide for 48 h successfully established a POI model， marked by a significant inhibition of KGN cell proliferation. Notably， the inhibitory effect of cyclophosphamide on KGN cell proliferation exhibited a positive correlation with its concentration. Zhuluan decoction-containing serum at 20% and 30% promoted cell proliferation and mitigated the inhibitory effect of cyclophosphamide on KGN cell proliferation， with comparable therapeutic efficacy observed at both concentrations. Compared with the blank control group， the model group displayed an elevated apoptosis rate （P<0.01）， reduced protein levels of PI3K， p-Akt， and p-mTOR （P<0.01）， increased protein levels of Beclin1， LC3， and ATG5 （P<0.01）， no significant alterations in the protein levels of Akt and mTOR， and an enhanced MDC autophagy fluorescence intensity （P<0.01）. In comparison to that the model group， the apoptosis rates in the blank control group， model group， Zhuluan decoction-containing serum group， autophagy inhibitor group， autophagy inhibitor + Zhuluan decoction-containing serum group， and estradiol valerate group all reduced （P<0.05， P<0.01）， with the most pronounced reduction observed in the autophagy inhibitor + Zhuluan decoction-containing serum group. The protein levels of PI3K， p-Akt， and p-mTOR were higher in other groups than in the model group （P<0.05， P<0.01）， being the highest in the autophagy inhibitor + Zhuluan decoctio-containing serum group （P<0.01）. The protein levels of Beclin1 and ATG5 were lower in other groups than in the model group （P<0.05， P<0.01）. The expression level of LC3 declined in the Zhuluan decoction-containing serum group and the estradiol valerate group （P<0.05， P<0.01）， while it decreased without statistical significance in the autophagy inhibitor group and the autophagy inhibitor + Zhuluan decoction-containing serum group. ConclusionZhuluan decoction may activate the PI3K/Akt/mTOR pathway to inhibit excessive autophagy and counteract the detrimental effects of cyclophosphamide on the KGN cell model， thus managing POI.

ObjectiveTo elucidate the underlying mechanism through which Zhuluan decoction suppresses excessive autophagy in human ovarian granulosa cells （KGN） and ameliorates premature ovarian insufficiency （POI） via the phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/mammalian target of rapamycin （mTOR） signaling pathway. MethodsThe optimal concentration of cyclophosphamide for inducing a POI model in KGN cells was identified via the cell counting kit-8 （CCK-8） assay. Subsequently， the impacts of varying concentrations of Zhuluan decoction-containing serum on the viability of the KGN cell model were assessed. After the optimal drug concentration was determined， KGN cells were categorized into the following groups： blank control （20% blank serum）， model （20% blank serum + 5 μmol·L^-1 cyclophosphamide）， Zhuluan decoction-containing serum （20% Zhuluan decoction-containing serum + 5 μmol·L^-1 cyclophosphamide）， autophagy inhibitor （20% blank serum + 5 μmol·L^-1 cyclophosphamide + 20 μmol·L^-1 chloroquine phosphate）， autophagy inhibitor + Zhuluan decoction-containing serum （20% Zhuluan decoction-containing serum + 5 μmol·L^-1 cyclophosphamide + 20 μmol·L^-1 chloroquine phosphate）， and estradiol valerate （20% estradiol valerate-containing serum + 5 μmol·L^-1 cyclophosphamide）. Following 48 hours of incubation， flow cytometry was utilized to measure the apoptosis rate of KGN cells in each group. Western blotting was employed to quantify the protein levels of PI3K， phosphorylated （p）-Akt， Akt， p-mTOR， and mTOR， along with the expression levels of autophagy-related proteins such as Beclin1， autophagy-related 5 homolog （ATG5）， and microtubule-associated protein 1 light chain 3 （LC3）， in each group. Additionally， monodansylcadaverine （MDC） staining was performed to evaluate the extent of autophagy in each group. ResultsIncubation of KGN cells with 5 μmol·L^-1 cyclophosphamide for 48 h successfully established a POI model， marked by a significant inhibition of KGN cell proliferation. Notably， the inhibitory effect of cyclophosphamide on KGN cell proliferation exhibited a positive correlation with its concentration. Zhuluan decoction-containing serum at 20% and 30% promoted cell proliferation and mitigated the inhibitory effect of cyclophosphamide on KGN cell proliferation， with comparable therapeutic efficacy observed at both concentrations. Compared with the blank control group， the model group displayed an elevated apoptosis rate （P<0.01）， reduced protein levels of PI3K， p-Akt， and p-mTOR （P<0.01）， increased protein levels of Beclin1， LC3， and ATG5 （P<0.01）， no significant alterations in the protein levels of Akt and mTOR， and an enhanced MDC autophagy fluorescence intensity （P<0.01）. In comparison to that the model group， the apoptosis rates in the blank control group， model group， Zhuluan decoction-containing serum group， autophagy inhibitor group， autophagy inhibitor + Zhuluan decoction-containing serum group， and estradiol valerate group all reduced （P<0.05， P<0.01）， with the most pronounced reduction observed in the autophagy inhibitor + Zhuluan decoction-containing serum group. The protein levels of PI3K， p-Akt， and p-mTOR were higher in other groups than in the model group （P<0.05， P<0.01）， being the highest in the autophagy inhibitor + Zhuluan decoctio-containing serum group （P<0.01）. The protein levels of Beclin1 and ATG5 were lower in other groups than in the model group （P<0.05， P<0.01）. The expression level of LC3 declined in the Zhuluan decoction-containing serum group and the estradiol valerate group （P<0.05， P<0.01）， while it decreased without statistical significance in the autophagy inhibitor group and the autophagy inhibitor + Zhuluan decoction-containing serum group. ConclusionZhuluan decoction may activate the PI3K/Akt/mTOR pathway to inhibit excessive autophagy and counteract the detrimental effects of cyclophosphamide on the KGN cell model， thus managing POI.

OBJECTIVE: To investigate the current status of clinical genetics specialization development and the diagnostic and therapeutic capabilities for hereditary diseases across medical institutions in Shanghai, and to assess the necessity and feasibility of establishing training bases for clinical genetics specialists. METHODS: By employing a cross-sectional survey design, the Clinical Genetics Committee of Shanghai Medical Association has conducted questionnaire surveys from March to April 2025 across 54 healthcare institutions in Shanghai (including 33 tertiary hospitals and 21 secondary hospitals). The survey involved administrative departments and medical personnel from 15 clinical specialties. The survey has covered current genetic disease diagnosis and treatment practices, relevant and specialised disease types, genetic department establishment, testing capabilities, personnel teams, and training requirements. RESULTS: The results revealed that 78.0% of clinical departments surveyed had treated patients with hereditary disorders. Shanghai possesses diagnostic and therapeutic expertise for over 95% of hereditary diseases listed in its rare disease catalogue, reflecting both the practical clinical demand for such conditions and the city's overall diagnostic and therapeutic strengths in this field. Nevertheless, significant disparities exist in the development of genetics departments across different tiers of healthcare institutions. Resources for genetic testing capabilities (including molecular, cellular, and biochemical testing) are also unevenly distributed across different tiers of hospitals. The survey further revealed that only 26.0% of departments believe that their current physician structure fully meets the diagnostic and treatment demands. Over 90% of departments consider standard training for clinical genetic specialists necessary, with 74.0% expressing willingness to participate in establishing training bases. Based on above findings and thorough deliberation, the Clinical Genetics Committee of the Shanghai Medical Association proposes advancing specialist training and discipline development through establishing a standard training system. The committee has drafted a three-year training protocol featuring a "joint training"-centered model, recommending a pilot-first, dynamically optimized strategy for steadily advancing training base development. CONCLUSION Shanghai faces substantial demand for genetic disease diagnosis and treatment, yet exhibits shortcomings in clinical genetics specialization development, resource allocation, and talent pipeline cultivation. To establish a standard training system holds significant practical importance and is underpinned by a broad demand.
Humans ; China ; Surveys and Questionnaires ; Genetic Diseases, Inborn/genetics* ; Cross-Sectional Studies ; Genetics, Medical/education* ; Genetic Testing

ObjectiveTo explore the mechanisms of Yangjing Tongluo prescription （YJTL） in the treatment of intrauterine adhesion （IUA） from the perspective of oxidative stress mediated by the Kelch-like ECH-associated protein 1/nuclear factor erythroid 2-related factor 2/heme oxygenase-1 （Keap1/Nrf2/HO-1） signaling pathway. MethodsA total of 48 rats with normal estrous cycles were selected and randomly divided into a normal group （n=8） and a modeling group （n=40）. An IUA rat model was established using a dual-injury method combining surgical curettage and infection. Eight rats were randomly selected from the modeling group for a pilot experiment to confirm successful model establishment. After successful modeling， the remaining 32 rats were randomly divided into a model group， a low-dose YJTL group （YJTL-L）， a high-dose YJTL group （YJTL-H）， and a Progynova group. Rats in the normal and model groups were administered purified water （15 mL·kg^-1） by gavage daily， while rats in the YJTL-L， YJTL-H， and Progynova groups received YJTL at doses of 6.43 and 12.86 g·kg^-1 and Progynova at 2.06 × 10^-4 g·kg^-1， respectively， for 14 consecutive days. The general condition， uterine morphology， and uterine index of the rats were monitored. Histopathological changes in uterine tissue were observed using hematoxylin-eosin （HE） staining. Serum levels of reactive oxygen species （ROS） and glutathione peroxidase （GSH-Px） were measured by enzyme-linked immunosorbent assay （ELISA）. Protein expression levels of Keap1， Nrf2， and HO-1 in endometrial tissue were detected by Western blot. Immunofluorescence （IF） was used to assess the distribution of Nrf2 and HO-1， as well as the expression of Nrf2 in the cytoplasm and nucleus. ResultsCompared with the normal group， rats in the model group exhibited poor mental status and reduced mobility， markedly edematous and tortuous uterine morphology， decreased gland number， and inflammatory reactions in the endometrium， along with an increased uterine organ index （P<0.05）. Serum ROS levels were significantly increased （P<0.05）， while serum GSH-Px levels were significantly decreased （P<0.05）. In endometrial tissue， Keap1 protein expression was increased （P<0.05）， whereas Nrf2 and HO-1 protein expression was decreased. Mild nuclear translocation of Nrf2 was observed， accompanied by increased relative fluorescence intensity of nuclear Nrf2 and decreased relative fluorescence intensity of cytoplasmic HO-1. Compared with the model group， all treatment groups showed varying degrees of improvement in the above symptoms and pathological changes. Serum ROS levels were reduced （P<0.05）， serum GSH-Px levels were increased （P<0.05）， Keap1 protein expression in endometrial tissue was decreased， and Nrf2 and HO-1 protein expression was increased in a dose-dependent manner （P<0.05）. Notably， significant nuclear translocation of Nrf2 was observed， with correspondingly increased relative fluorescence intensity of nuclear Nrf2 and enhanced relative fluorescence intensity of cytoplasmic HO-1. ConclusionYJTL may enhance antioxidant capacity and repair oxidative damage to the endometrial basal layer by regulating the Keap1/Nrf2/HO-1 signaling pathway.

Based on spleen deficiency-phlegm dampness as the core pathogenesis of obesity， and integrating recent advances in modern medicine regarding the key role of immune inflammation in obesity， this paper proposes a multidimensional pathogenic network of "obesity-spleen deficiency-phlegm dampness-immune imbalance". Various traditional Chinese medicine （TCM） herbs that strengthen the spleen， regulate qi， and resolve phlegm and dampness can treat obesity by improving spleen-stomach transport and transformation， promoting water-damp metabolism， and regulating immune homeostasis. This highlights immune inflammation as an important entry point to elucidate the TCM concepts of "spleen deficiency-phlegm dampness" and the therapeutic principle of "strengthening the spleen and eliminating dampness to treat obesity". By systematically analyzing the intrinsic connection between "spleen deficiency generating dampness， internal accumulation of phlegm dampness" and immune dysregulation in obesity， this paper aims to provide theoretical support for TCM treatment of obesity based on dampness.

ObjectiveTo investigate the mechanism by which Zhuluan decoction improves ovarian function in a cyclophosphamide （CTX）-induced premature ovarian insufficiency （POI） rat model through regulating the adenosine monophosphate-activated protein kinase （AMPK）/mammalian target of rapamycin （mTOR）/Unc-51-like kinase 1 （ULK1） signaling pathway. MethodsA POI rat model was established via intraperitoneal injection of CTX. Ten rats were randomly selected as the normal control group. The remaining 50 successfully modeled rats were randomly divided into the model group， the estradiol valerate group （0.1 mg·kg^-1）， low-dose （7 g·kg^-1）， medium-dose （14 g·kg^-1）， and high-dose （28 g·kg^-1） Zhuluan decoction groups， with 10 rats per group. All groups received intragastric administration for 21 consecutive days. Changes in body weight， uterine， and ovarian indices were observed across groups. Hematoxylin and eosin （HE） staining was used to examine ovarian pathological alterations and count follicles at various stages in cross-sections. Enzyme-linked immunosorbent assay （ELISA） measured serum estradiol （E₂）， follicle-stimulating hormone （FSH）， and anti-Müllerian hormone （AMH） levels. Western blot （WB） and immunohistochemistry （IHC） were used to detect the expression levels of AMPK， mTOR， ULK1 and their phosphorylated proteins， Beclin1， Bcl-2-associated X protein （Bax）， and Bcl-2 in ovarian tissues of each group. ResultsCompared with the normal group， model rats exhibited slower body weight gain （P<0.01）， significantly reduced uterine and ovarian organ indices （P<0.01）， reduced ovarian volume， irregular follicular morphology， unclear cumulus and oocyte structures， and a significant reduction in the number of primordial follicles （P<0.01）. Conversely，the numbers of secondary and atretic follicles significantly increased （P<0.01）. Serum E₂ and AMH levels significantly decreased， while FSH levels significantly elevated（P<0.01）. The expression of AMPK， ULK1， Bax， and Beclin1 was increased （P<0.01）， while that of mTOR and Bcl-2 was decreased （P<0.01）. Compared with the model group， the estradiol valerate group and the medium- and high-dose Zhuluan decoction groups showed significantly increased body weight， uterine organ index， and ovarian organ index （P<0.01）， while the low-dose Zhuluan decoction group showed no statistically significant difference. Ovarian volume increased across all treatment groups， with improved follicular morphology and structure. The number of primordial follicles significantly increased （P<0.01）， while the number of atretic follicles significantly decreased （P<0.01）. The number of secondary follicles decreased in the estradiol valerate group and the high-dose Zhuluan decoction group （P<0.01）， with no significant difference in the low- and medium-dose Zhuluan decoction groups. E₂ and AMH levels significantly increased， while FSH levels significantly decreased in all treatment groups （P<0.05， P<0.01）. The estradiol valerate group and medium- and high-dose Zhuluan decoction groups showed significantly reduced expression of AMPK， ULK1， Bax， and Beclin1 （P<0.05， P<0.01）， while mTOR and Bcl-2 expression levels were significantly elevated （P<0.01）. The low-dose Zhuluan decoction group exhibited significantly reduced expression of ULK1， Beclin1， and Bax （P<0.05）， while Bcl-2 expression was significantly increased （P<0.05）. ConclusionZhuluan decoction improves serum hormone levels in POI rats and mitigates chemotherapy （CTX）-induced damage to ovarian structure and function， exerting ovarian protective effect. This mechanism may involve regulating the AMPK/mTOR/ULK1 signaling pathway to inhibit excessive autophagy and apoptosis.

ObjectiveTo explore the effects of electroacupuncture at governor vessel or Jiaji acupoints on the activation of lumbar spinal cord neurons and the recovery of motor function in paralyzed hind limbs in rats with complete transection of spinal cord injury. MethodsTwenty adult female SD rats were selected. Among them， 15 rats underwent the complete transection of the 10th thoracic spinal cord and were randomly divided into the Control group， the electroacupuncture at governor vessel acupoints （GV） group and the electroacupuncture at Jiaji acupoints （JJ） group， with 5 rats in each group. The other 5 rats served as the Normal group. Electroacupuncture treatment was initiated 3 days after surgery in the GV and JJ groups and performed every other day. Basso-Beatti-Bresnahan （BBB） behavioral score was evaluated weekly. Eight weeks after the operation， cortical motor evoked potential （CMEP） detection was performed， followed by perfusion fixation， tissue harvest， frozen sectioning， and immunofluorescence staining experiments. Results① Both electroacupuncture treatments activated the expression of the excitatory marker c-Fos in L1 spinal cord neurons. The c-Fos⁺ activated neurons were mainly distributed in lamina Ⅳ， Ⅴ and Ⅶ of the L1 spinal cord， while no c-Fos⁺ neuron was observed in the Normal group and the Control group. The percentages of c-Fos⁺ neurons activated by electroacupuncture in the GV group and the JJ group were （10.23±0.15） % and （10.77±1.12） %， respectively， and there was no significant difference between the two electroacupuncture groups. ② vGluT1⁺ proprioceptive afferent nerve fibers were mainly distributed in the dorsal horn and lamina Ⅶ of intermediate zone of L1 spinal cord gray matter. In the dorsal horn of L1 spinal cord， the relative fluorescence intensity of vGluT1⁺ nerve fibers in the GV group and the JJ group were greater than that in the Control group， and that in the JJ group was greater than that in the GV group. In the lamina Ⅶ in intermediate zone of L1 spinal cord， where the central pattern generator （CPG） for hindlimb rhythmic movement is located， the relative fluorescence intensity of vGluT1⁺ nerve fibers in the GV group and the JJ group were significantly higher than that in the Control group， and the JJ group was also higher than the GV group； The lamina Ⅶ neurons innervated by vGluT1⁺ axon terminals were activated to express the excitatory marker c-Fos in the GV group and the JJ group. ③ The immunofluorescence staining results of acetylcholinesterase （ChAT） in the ventral horn of L4 spinal cord of each group showed that the number of surviving ChAT⁺ motor neurons in the GV group （19.1±3.4， P=0.000 2） and JJ group （19.0±2.9， P=0.000 2） were higher than that in the Control group （9.9±1.9）. ④ The quantitative analysis of the vGluT2⁺ excitatory axonal terminals， vGAT⁺ inhibitory axonal terminals， and the ratio of vGluT2⁺/vGAT⁺ axonal terminals ［reflecting the excitation/inhibition （E/I） balance of motor neurons］ innervating ChAT⁺ motor neurons in the ventral horn of the L4 spinal cord revealed that all three indices were significantly higher in the GV and JJ groups than in the Control group. Moreover， the E/I ratio of motor neurons in the two electroacupuncture groups gradually reached the same level as normal and showed no statistical difference from the Normal group. ⑤ The electrophysiological and behavioral detection results showed that compared with the Control group， the latencies of CMEP in the GV group and the JJ group shortened， while their amplitudes increased. The BBB score of paralyzed hindlimbs showed that after 8 weeks of treatment， the scores of both the GV group （4.25±1.26， P=0.023 4） and the JJ group （4.75±0.96， P=0.000 2） were higher than those of the Control group （1.80±1.30）， but there was no statistical difference between the two electroacupuncture groups. ConclusionElectroacupuncture at GV or JJ acupoints may promote the afferent of vGluT1⁺ proprioceptive nerve fibers into the spinal cord， indicating the possibility to activate CPG neurons in lamina Ⅶ of L1 spinal cord； facilitate the survival of motor neurons in the L4 spinal cord and restore their excitatory/inhibitory balance， improving the motor function of the paralyzed hindlimbs in rats.

ObjectiveTo investigate the mechanism by which Zhuluan decoction improves ovarian function in a cyclophosphamide （CTX）-induced premature ovarian insufficiency （POI） rat model through regulating the adenosine monophosphate-activated protein kinase （AMPK）/mammalian target of rapamycin （mTOR）/Unc-51-like kinase 1 （ULK1） signaling pathway. MethodsA POI rat model was established via intraperitoneal injection of CTX. Ten rats were randomly selected as the normal control group. The remaining 50 successfully modeled rats were randomly divided into the model group， the estradiol valerate group （0.1 mg·kg^-1）， low-dose （7 g·kg^-1）， medium-dose （14 g·kg^-1）， and high-dose （28 g·kg^-1） Zhuluan decoction groups， with 10 rats per group. All groups received intragastric administration for 21 consecutive days. Changes in body weight， uterine， and ovarian indices were observed across groups. Hematoxylin and eosin （HE） staining was used to examine ovarian pathological alterations and count follicles at various stages in cross-sections. Enzyme-linked immunosorbent assay （ELISA） measured serum estradiol （E₂）， follicle-stimulating hormone （FSH）， and anti-Müllerian hormone （AMH） levels. Western blot （WB） and immunohistochemistry （IHC） were used to detect the expression levels of AMPK， mTOR， ULK1 and their phosphorylated proteins， Beclin1， Bcl-2-associated X protein （Bax）， and Bcl-2 in ovarian tissues of each group. ResultsCompared with the normal group， model rats exhibited slower body weight gain （P<0.01）， significantly reduced uterine and ovarian organ indices （P<0.01）， reduced ovarian volume， irregular follicular morphology， unclear cumulus and oocyte structures， and a significant reduction in the number of primordial follicles （P<0.01）. Conversely，the numbers of secondary and atretic follicles significantly increased （P<0.01）. Serum E₂ and AMH levels significantly decreased， while FSH levels significantly elevated（P<0.01）. The expression of AMPK， ULK1， Bax， and Beclin1 was increased （P<0.01）， while that of mTOR and Bcl-2 was decreased （P<0.01）. Compared with the model group， the estradiol valerate group and the medium- and high-dose Zhuluan decoction groups showed significantly increased body weight， uterine organ index， and ovarian organ index （P<0.01）， while the low-dose Zhuluan decoction group showed no statistically significant difference. Ovarian volume increased across all treatment groups， with improved follicular morphology and structure. The number of primordial follicles significantly increased （P<0.01）， while the number of atretic follicles significantly decreased （P<0.01）. The number of secondary follicles decreased in the estradiol valerate group and the high-dose Zhuluan decoction group （P<0.01）， with no significant difference in the low- and medium-dose Zhuluan decoction groups. E₂ and AMH levels significantly increased， while FSH levels significantly decreased in all treatment groups （P<0.05， P<0.01）. The estradiol valerate group and medium- and high-dose Zhuluan decoction groups showed significantly reduced expression of AMPK， ULK1， Bax， and Beclin1 （P<0.05， P<0.01）， while mTOR and Bcl-2 expression levels were significantly elevated （P<0.01）. The low-dose Zhuluan decoction group exhibited significantly reduced expression of ULK1， Beclin1， and Bax （P<0.05）， while Bcl-2 expression was significantly increased （P<0.05）. ConclusionZhuluan decoction improves serum hormone levels in POI rats and mitigates chemotherapy （CTX）-induced damage to ovarian structure and function， exerting ovarian protective effect. This mechanism may involve regulating the AMPK/mTOR/ULK1 signaling pathway to inhibit excessive autophagy and apoptosis.