Objective:To investigate the mechanism of recombinant human mesencephalic astrocyte-derived neurotrophic factor (rhMANF) in spinal cord injury (SCI) repair promoted by A2 reactive astrocyte polarization.Methods:One hundred and twenty female SPF SD rats were randomly divided into sham-operated group, SCI group, SCI+control group and SCI+rhMANF group ( n=30 in each group). SCI models were prepared by heavy drop method in the later 3 groups, and 10 μL sterile saline or 10 μL sterile saline+5 μg rhMANF were injected intrathecally in the later 2 groups 30 min after modeling. Basso-Beattie-Bresnahan (BBB) scale was used to evaluate the motor function in each group 1, 3, 7, 14, 21 and 28 days after injection. After behavioral assessment 3 days after injection, the protein expressions of ReIB, p52, phosphorylated (p)-ReIB and p-p52 in the spinal cord tissues were detected by Western blotting, and the expressions of anti-inflammatory cytokine and neurotrophic factor in the spinal cord tissues were detected by ELISA. After behavioral assessment 14 days after injection, immunofluorescent staining was performed to detect the expressions of neuronal nuclear antigen (NeuN), Syn and S100A10 in the spinal cord tissues. After behavioral assessment 28 days after injection, HE staining and uranyl acetate-lead citrate double staining were used to observe the pathological changes of the spinal cord under light microscope and electron microscope, respectively. Results:On 14, 21, and 28 days after injection, the BBB score in the SCI+rhMANF group was significantly higher than that in the SCI group and SCI+control group ( P<0.05). On 3 days after injection, the p-ReiB and p-p52 protein expressions in the SCI+rhMANF group (1.17±0.02 and 1.00±0.07) were significantly higher than those in the SCI group (0.74±0.01 and 0.42±0.11) and SCI+control group (0.79±0.00 and 0.64±0.02, P<0.05); the SCI+rhMANF group had significantly increased interleukin (IL)-4, IL-10, IL-13, neurotrophin-3, transforming growth factor-β and granulocyte colony-stimulating factor expressions ([217.58±16.06] pg/mg, [276.53±15.00]) pg/mg, [178.88±7.03] pg/mg, [172.61±16.43] pg/mg, [241.00±15.80] pg/mg, and [166.63±14.61] pg/mg) compared with the SCI group ([132.15±18.86] pg/mg, [173.48±18.24] pg/mg, [109.01±3.79] pg/mg, [104.64±18.21] pg/mg, [138.09±9.93] pg/mg, and [91.26±11.09] pg/mg), and SCI+control group ([137.80±27.70] pg/mg, [185.78±19.20] pg/mg, [112.44±13.51] pg/mg, [93.13±22.09] pg/mg, [159.48±32.50] pg/mg, and [112.67±18.32] pg/mg, P<0.05). On 14 days after injection, the immunofluorescent staining intensities of NeuN/S100A10, NeuN/Syn in the SCI+rhMANF group (2.51±0.24/2.85±0.27 and 2.48±0.35/1.92±0.32) were significantly higher than those in the SCI group (0.99±0.11/1.00±0.18 and 1.00±0.19/1.00±0.08) and SCI+control group (1.39±0.09/0.93±0.20 and 1.26±0.35/0.94±0.19, P<0.05). Light microscopy showed that the spinal cord nerve tissues in the SCI group and SCI+control group had loose structure, with edema and vacuolar degeneration; those in the sham-operated group and SCI+rhMANF group had dense structure, with round and cone-shaped neurons and large and round nucleus, and without vacuolar degeneration. Transmission electron microscopy showed intact structure of myelin sheath and axon in the sham-operated group, loose and shrunked spinal cord nerve cells (chromatin condensation, and cell membrane bleb formation) in the SCI group and SCI+control group, and relatively complete cell structure in the SCI+rhMANF group. Conclusion:The rhMANF can activate ReIB/P52 nuclear translocation phosphorylation, up-regulate the anti-inflammatory factor and neurotrophic factor expressions, induce the A2 astrocyte polarization, and promote the synaptic growth and spinal cord injury recovery.

BACKGROUND:The pathogenesis of diabetic peripheral neuropathy has not yet been clarified,and TAM(Tyro3,Axl,and MerTK)receptor tyrosine kinases can control apoptotic cells and suppress inflammatory responses in the central nervous system. OBJECTIVE:To investigate the difference of Mer receptor tyrosine kinase(MerTK)levels in plasma and sciatic nerve tissue of Sprague-Dawley rats with type 2 diabetes and diabetic peripheral neuropathy,and to study the correlation between MerTK and diabetic peripheral neuropathy. METHODS:Forty male Sprague-Dawley were randomly divided into control group with 15 rats,type 2 diabetes group with 10 rats,and diabetic peripheral neuropathy group with 15 rats.The control group was fed with ordinary diet,while the experimental groups were fed with high-fat and high-sugar diet.After 6 weeks,intraperitoneal injection of streptozotocin at the minimum dose of 35 mg/kg was administered in the two experimental groups.After 14 days,tail vein blood was collected to detect blood glucose.If blood glucose≥16.7 mmol/L,the model of type 2 diabetes was successfully established.Rats in the diabetic peripheral neuropathy group continued to be fed with a high-sugar and high-fat diet for 8 weeks.The sciatic nerve conduction velocity of rats was detected through live isolation under anesthesia.Blood samples were collected from the abdominal aorta,and the sciatic nerve tissue was collected.Histological changes of nerve fibers in each group were observed under a light microscope to confirm the success of diabetic peripheral neuropathy modeling.ELISA was used to detect peripheral blood glucose,blood lipids and serum MerTK levels in rats;hematoxylin-eosin staining was used to observe the histological changes in the sciatic nerve;immunofluorescence,immunohistochemistry and western blot were used to detect the expression of MerTK in the sciatic nerve tissue. RESULTS AND CONCLUSION:The Sprague-Dawley rat models of type 2 diabetes and type 2 diabetes peripheral neuropathy were successfully constructed,and the modeling rate of diabetic peripheral neuropathy was 80%.Compared with the control group,the blood glucose levels of rats in the type 2 diabetes and diabetic peripheral neuropathy groups were significantly higher(P<0.000 1),while the blood glucose level in the diabetic peripheral neuropathy group was higher than that in the type 2 diabetes group;and the sciatic nerve conduction velocity was significantly decreased(P<0.05),which was lower in the diabetic peripheral neuropathy group than the type 2 diabetes group.Histological examination:Compared with the control group,the sciatic nerve nuclei were reduced in the type 2 diabetes group,with some vacuolar degeneration and phagocytosis;in the diabetic peripheral neuropathy group,the cell body was swollen,the nuclear spacing was increased,vacuolar degeneration was observed,and the myelin sheath was partitioned and unsmooth,and lattice-like axons appeared.Serum MerTK levels were significantly higher in the diabetic peripheral neuropathy group than the control group.Expression of MerTK in the sciatic nerve tissue was significantly upregulated in the diabetic peripheral neuropathy group compared with the control group(P<0.05).To conclude,elevated levels of MerTK in plasma and sciatic nerve tissue of rats with diabetic peripheral neuropathy are presumably related to its anti-inflammatory and immunomodulatory effects.

Chlorobenzene contaminants (CBs) pose a threat to the eco-environment, and functional strains hold considerable potential for the remediation of CB-contaminated sites. To deeply explore the application potential of functional bacteria in the in-situ bioremediation of CBs, this study focused on the biodegradation characteristics and degradation kinetics of CB and 1, 2-dichlorobenzene (1, 2-DCB) in soil by the isolated strain Serratia marcescens TF-1. Additionally, an in-situ remediation trial was conducted with this strain at a chemical industrial site. Batch serum bottle experiments showed that the degradation rate of CB at the concentrations ranging from 20 to 200 mg/L by TF-1 was 0.22-0.66 mol/(g_cell·h), following the Haldane model, with the optimal concentration at 23.12 mg/L. The results from simulated soil degradation experiments indicated that the combined use of TF-1 and sodium succinate (SS) significantly enhanced the degradation of CBs, with the maximum degradation rate of CB reaching 0.104 d^-1 and a half-life of 6.66 d. For 1, 2-DCB, the maximum degradation rate constant was 0.068 7 d^-1, with a half-life of 10.087 d. The in-situ remediation results at the chemically contaminated site demonstrated that the introduction of bacterial inoculant and SS significantly improved the removal of CBs, achieving the removal rates of 84.2%-100% after 10 d. CB, 1, 4-dichlorobenzene (1, 4-DCB), and benzo[a]pyrene were completely removed. Microbial diversity analysis revealed that the in-situ remediation facilitated the colonization of TF-1 and the enrichment of indigenous nitrogen-fixing Azoarcus, which may have played a key role in the degradation process. This study provides a theoretical basis and practical experience for the in situ bioremediation of CBs-contaminated sites.
Chlorobenzenes/isolation & purification* ; Biodegradation, Environmental ; Soil Pollutants/isolation & purification* ; Serratia marcescens/metabolism* ; Industrial Waste ; Soil Microbiology

Objective: To investigate the potential efficacy and safety of Lutai Danshen Baishao granules (LDBG) for treating female melasma associated with kidney deficiency and blood stasis patterns. Methods: A randomized, double-blind, placebo-controlled trial was conducted at the Third Central Hospital of Tianjin, China from March to December 2023. A total of 110 female patients with melasma linked to kidney deficiency and blood stasis were enrolled and treated with either LDBG or a placebo twice daily for 60 days. Efficacy was assessed through measures such as the total melasma area, reduced melasma area, reduction rate of melasma area, melasma color score, Melasma Area and Severity Index (MASI) score, and traditional Chinese medicine (TCM) symptom score scale. Safety assessments included routine blood and biochemical tests. Results: Participants in both groups were aged 52–63 years, with no significant differences. After the 2-month intervention, the total melasma area decreased in both groups; however, a greater reduction was observed in the test group [462.50 mm2 (12.81%) vs. 100.00 mm2 (3.11%), P < .001]. Moreover, LDBG treatment significantly reduced the MASI and melasma color scores in the test group (P < .05). The total TCM symptom evaluation score significantly decreased (test group: 6.00 vs. placebo group: 7.00, P = .001), with significant relief in symptoms such as improvement in dark lips, nails, and waist soreness in the test group, compared with that in the placebo group (P < .05). Within-group comparisons revealed that TCM syndrome was significantly alleviated in the test group (P < .05). Conclusion LDBG intervention shows promising effectiveness in reducing female melasma and alleviating TCM syndromes.

Objective:To investigate the mechanism of recombinant human mesencephalic astrocyte-derived neurotrophic factor (rhMANF) in spinal cord injury (SCI) repair promoted by A2 reactive astrocyte polarization.Methods:One hundred and twenty female SPF SD rats were randomly divided into sham-operated group, SCI group, SCI+control group and SCI+rhMANF group ( n=30 in each group). SCI models were prepared by heavy drop method in the later 3 groups, and 10 μL sterile saline or 10 μL sterile saline+5 μg rhMANF were injected intrathecally in the later 2 groups 30 min after modeling. Basso-Beattie-Bresnahan (BBB) scale was used to evaluate the motor function in each group 1, 3, 7, 14, 21 and 28 days after injection. After behavioral assessment 3 days after injection, the protein expressions of ReIB, p52, phosphorylated (p)-ReIB and p-p52 in the spinal cord tissues were detected by Western blotting, and the expressions of anti-inflammatory cytokine and neurotrophic factor in the spinal cord tissues were detected by ELISA. After behavioral assessment 14 days after injection, immunofluorescent staining was performed to detect the expressions of neuronal nuclear antigen (NeuN), Syn and S100A10 in the spinal cord tissues. After behavioral assessment 28 days after injection, HE staining and uranyl acetate-lead citrate double staining were used to observe the pathological changes of the spinal cord under light microscope and electron microscope, respectively. Results:On 14, 21, and 28 days after injection, the BBB score in the SCI+rhMANF group was significantly higher than that in the SCI group and SCI+control group ( P<0.05). On 3 days after injection, the p-ReiB and p-p52 protein expressions in the SCI+rhMANF group (1.17±0.02 and 1.00±0.07) were significantly higher than those in the SCI group (0.74±0.01 and 0.42±0.11) and SCI+control group (0.79±0.00 and 0.64±0.02, P<0.05); the SCI+rhMANF group had significantly increased interleukin (IL)-4, IL-10, IL-13, neurotrophin-3, transforming growth factor-β and granulocyte colony-stimulating factor expressions ([217.58±16.06] pg/mg, [276.53±15.00]) pg/mg, [178.88±7.03] pg/mg, [172.61±16.43] pg/mg, [241.00±15.80] pg/mg, and [166.63±14.61] pg/mg) compared with the SCI group ([132.15±18.86] pg/mg, [173.48±18.24] pg/mg, [109.01±3.79] pg/mg, [104.64±18.21] pg/mg, [138.09±9.93] pg/mg, and [91.26±11.09] pg/mg), and SCI+control group ([137.80±27.70] pg/mg, [185.78±19.20] pg/mg, [112.44±13.51] pg/mg, [93.13±22.09] pg/mg, [159.48±32.50] pg/mg, and [112.67±18.32] pg/mg, P<0.05). On 14 days after injection, the immunofluorescent staining intensities of NeuN/S100A10, NeuN/Syn in the SCI+rhMANF group (2.51±0.24/2.85±0.27 and 2.48±0.35/1.92±0.32) were significantly higher than those in the SCI group (0.99±0.11/1.00±0.18 and 1.00±0.19/1.00±0.08) and SCI+control group (1.39±0.09/0.93±0.20 and 1.26±0.35/0.94±0.19, P<0.05). Light microscopy showed that the spinal cord nerve tissues in the SCI group and SCI+control group had loose structure, with edema and vacuolar degeneration; those in the sham-operated group and SCI+rhMANF group had dense structure, with round and cone-shaped neurons and large and round nucleus, and without vacuolar degeneration. Transmission electron microscopy showed intact structure of myelin sheath and axon in the sham-operated group, loose and shrunked spinal cord nerve cells (chromatin condensation, and cell membrane bleb formation) in the SCI group and SCI+control group, and relatively complete cell structure in the SCI+rhMANF group. Conclusion:The rhMANF can activate ReIB/P52 nuclear translocation phosphorylation, up-regulate the anti-inflammatory factor and neurotrophic factor expressions, induce the A2 astrocyte polarization, and promote the synaptic growth and spinal cord injury recovery.

OBJECTIVE To explore the effect of volatile oil of Ligusticum chuanxiong on the transdermal properties and cytotoxicity of triptolide in vitro. METHODS The chemical constituents of the volatile oil of L. chuanxiong were analyzed by gas chromatography-mass spectrometry. The lower abdominal skin of KM mice was separated and divided into triptolide group， triptolide in compatibility with volatile oil of L. chuanxiong groups at 1∶10， 1∶50， 1∶100 （hereinafter referred to as “compatibility 1∶10”“compatibility 1∶50”“compatibility 1∶100” groups）. After the skin of mice in each group was fully exposed to 0.2 g of the corresponding cream for 24 h， the cumulative transdermal dose （Qn） of triptolide in the receiving solution of each group was determined by high-performance liquid chromatography， and the transdermal absorption rate （Jss） was calculated. Human immortalized keratinocytes （HaCat） were used as a model， the CCK-8 method was used to detect the cell survival rate of different concentrations of the volatile oil of L. chuanxiong and triptolide before and after compatibility. RESULTS A total of 62 chemical constituents of the volatile oil of L. chuanxiong were identified， including Z-ligustilide， senkyunolide， and β-selinene. The Qn （P＜ 0.01） and Jss of triptolide increased within 24 h in the compatibility 1∶10 and 1∶50 groups， while the Qn （P＜0.05） and Jss decreased in the compatibility 1∶100 group as compared with the triptolide group. Compared with the triptolide group， the cell survival rate of HaCat was significantly increased in the compatibility 1∶10 and 1∶50 groups when the triptolide concentrations were 36， 72 and 144 ng/mL （P＜0.05 or P＜0.01）； while the cell survival rate of HaCat was decreased in the compatibility 1∶100 group， but the difference was not statistically significant （P＞0.05）. CONCLUSIONS When the compatibility ratio of triptolide and volatile oil of L. chuanxiong was 1∶10 or 1∶50， it can promote the transdermal absorption of triptolide and reduce the cytotoxicity of triptolide to HaCat.

Diabetic ulcer (DU), one of the common and serious complications in patients with diabetes mellitus, often leads to infection, necrosis and amputation, and has a long and costly treatment period. Because of DU's unclear healing mechanism and the difficulty of delayed healing, its treatment and management have been a major challenge in clinical medicine. In recent years, the potential role of mitochondrial autophagy in DU has become a research hotspot with the in-depth study of mitochondrial autophagy mechanism. Previous studies have shown that mitochondrial autophagy is an important intracellular self-repair mechanism that plays a crucial role in maintaining cellular health and functional stability. During the development of DU, mitochondrial autophagy plays multiple roles in attenuating oxidative stress and inflammatory responses, maintaining mitochondrial functional homeostasis, influencing cell proliferation and repair capacity during DU healing, promoting DU healing, and enhancing antimicrobial capacity. In this paper, we illustrate the multiple roles played by mitochondrial autophagy in DU prevention and treatment, as well as the potential applications of mitochondrial autophagy in DU therapy. It is expected to provide a basis for the clinical application of mitochondrial autophagy in DU treatment, and provide more effective strategies and solutions for the treatment of DU.

Diabetic ulcer (DU), one of the common and serious complications in patients with diabetes mellitus, often leads to infection, necrosis and amputation, and has a long and costly treatment period. Because of DU's unclear healing mechanism and the difficulty of delayed healing, its treatment and management have been a major challenge in clinical medicine. In recent years, the potential role of mitochondrial autophagy in DU has become a research hotspot with the in-depth study of mitochondrial autophagy mechanism. Previous studies have shown that mitochondrial autophagy is an important intracellular self-repair mechanism that plays a crucial role in maintaining cellular health and functional stability. During the development of DU, mitochondrial autophagy plays multiple roles in attenuating oxidative stress and inflammatory responses, maintaining mitochondrial functional homeostasis, influencing cell proliferation and repair capacity during DU healing, promoting DU healing, and enhancing antimicrobial capacity. In this paper, we illustrate the multiple roles played by mitochondrial autophagy in DU prevention and treatment, as well as the potential applications of mitochondrial autophagy in DU therapy. It is expected to provide a basis for the clinical application of mitochondrial autophagy in DU treatment, and provide more effective strategies and solutions for the treatment of DU.

Objective:To study the effects of electron beam irradiation and 60Co irradiation on the composition changes of four alkaloids in Sophorae Flavescentis Radix, intermediate extracts of Sophorae Flavescentis Radix and Lixieling Tablets. Methods:Sophorae Flavescentis Radix, intermediate extracts of Sophorae Flavescentis Radix and Lixieling Tablets were irradiated at different doses of 0, 1.5, 3, 5, 7, 10, 20, 30, 40 kGy by electron beam irradiation and 60Co irradiation. The contents of oxymatrine, oxysophocarpine, matrine and sophocarpine were determined by HPLC, and the changes of the components before and after irradiation were compared. Results:Oxymatrine, oxysophocarpine, matrine and sophocarpine were among 0.046 9-0.937 4 μg, 0.020 5-0.410 4 μg, 0.098 9-1.977 9 μg, 0.048 7-0.973 1 μg, respectively. The linear relationship was good. The average recovery rates were 98.1%, 100.1%, 100.5%, 96.6%, respectively, and the RSDs were 1.69%, 2.03%, 3.14% and 1.10%, respectively. Electron beam irradiation and 60Co irradiation had no statistical significance on the changes of oxymatrine, oxysophocarpine, matrine and sophocarpine in Sophora flavescens, but had statistical significance in the contents of intermediate extracts of Sophorae Flavescentis Radix and Lixieling Tablets. Conclusion:The established method for the determination of matrine is accurate, reproducible, simple and practical, and can be used for the quality control of Lixieling Tablets. Irradiation has no significant effect on the content of Sophorae Flavescentis Radix, while high dose irradiation has significant effect on the intermediates and finished products of Sophorae Flavescentis Radix, which can provide a basis for quality control and sterilization irradiation of enterprises.

Objective To explore the regulatory effect of Mertk expression level on NF-κb pathway in rat Schwann cells and its possible mechanism.Methods Rat Schwann cells were cultured in vitro,and the expression of Mertk in Schwann cells exposed to high glucose was detected by Western blot.Co-immunoprecipitation was used to detect the interaction between endogenous Mertk and Ikbkb.Western blot was used to detect the expression levels of Ikbkb,P65 and tumor necrosis factor-α in Schwann cells after Mertk silencing.The protein expressions of Mertk,Ikbkb and P65 after silencing Mertk were detected by immunofluorescence.Results Mertk was expressed in Schwann cells,and the expression level increased with the increase of glucose concentration.Co-immunoprecipitation assay showed that Mertk interacted with Ikbkb in rat Schwann cells.Compared with the control group,the expression level of Mertk was significantly decreased(P<0.05),while Ikbkb,P65 and TNF-α were significantly increased(P<0.05)after knock down expression of Mertk in Schwann cells.Immunofluorescence experiments showed that the fluorescence of Mertk was decreased,and the fluorescence of Ikbkb and P65 was increased in the silenced Schwann cells.Conclusion After the expression of Mertk is decreased,it can mediate the regulation of NF-κb pathway in Schwann cells through interaction with Ikbkb,and up-regulate the expression of P65 and inflammatory factor TNF-α.