ObjectiveBased on transcriptomics， to explore the mechanism of Hei Xiaoyaosan regulating the phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） signaling pathway to improve the learning and memory ability of insomnia rats with liver depression syndrome. MethodsSixty 8-week-old male SD rats were randomly divided into the blank group， model group， eszopiclone group （0.09 mg·kg^-1）， and low， medium， and high dose groups of Hei Xiaoyaosan （3.82， 7.65， 15.30 g·kg^-1）， with ten rats in each group. Except for the blank group， the other groups were induced insomnia rat model with liver depression by chronic restraint， tail clamping stimulation and intraperitoneal injection of p-chlorophenylalanine （PCPA）. Each treatment group received intragastric administration according to the specified dosage， once a day for 14 consecutive days. The pentobarbital sodium cooperative sleep test， open field test， and Morris water maze test were used to test the sleep quality， depressive-like behavior， and learning and memory abilities of rats. Additionally， enzyme-linked immunosorbent assay （ELISA） was used to detect the contents of 5-hydroxytryptamine （5-HT）， γ-aminobutyric acid （GABA）， brain-derived neurotrophic factor （BDNF）， glial cell line-derived neurotrophic factor （GDNF） and nitric oxide （NO） in hippocampus. Hematoxylin-eosin （HE） staining was performed to observe pathological changes of the hippocampal tissue， while terminal deoxynucleotidyl transferase deoxyuridine triphosphate （dUTP） nick end labeling （TUNEL） was used to evaluate apoptosis of hippocampal neurons. Transcriptomic sequencing technology was employed to identify differentially expressed genes in hippocampus between the model group and the blank group， as well as between the medium-dose group of Hei Xiaoyaosan and the model group. Gene Ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） enrichment analysis were performed on the intersecting genes. Subsequently， the enriched key genes and signaling pathways were analyzed and verified. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was utilized to assess the mRNA expression levels of phosphatase and tensin homolog （PTEN）， B-cell lymphoma-2 （Bcl-2）-like protein 11 （BCL2L11）， and mitogen-activated protein kinase 1 （MAPK1） in hippocampus， and Western blot was employed to evaluate the protein expressions of PI3K， phosphorylation （p）-PI3K， Akt， p-Akt， Bcl-2， Bcl-2-associated X protein （Bax）， and cleaved Caspase-3 in the same tissue. ResultsCompared with the blank group， the model group exhibited a reduction in body weight， an increase in sleep latency， and a decrease in sleep duration （P<0.01）. Additionally， rats showed obvious depression-like behavior， and their learning and memory abilities decreased. Furthermore， the contents of 5-HT， GABA， NO， BDNF and GDNF in hippocampus decreased （P<0.01）. Histological examination revealed a disorganized cell arrangement in the CA1 region of the hippocampus， characterized by irregular cell shapes， a reduced cell count， deeply stained and pyknotic nuclei， increased vacuolar degeneration， and an elevated apoptosis rate （P<0.01）. Compared with the model group， the body weight of the high and medium dose groups of Hei Xiaoyaosan increased， the sleep latency shortened and the sleep time prolonged （P<0.05， P<0.01）. Additionally， depression-like behavior and learning and memory abilities of rats were significantly improved， the levels of 5-HT， GABA， NO， BDNF and GDNF in the hippocampus increased （P<0.05， P<0.01）. These interventions also ameliorated pathological damage in the hippocampal CA1 area and reduced the apoptosis of hippocampal neurons （P<0.01）. Transcriptomic sequencing results indicated that Hei Xiaoyaosan might exert a therapeutic effect by regulating PI3K/Akt pathway through key mRNAs such as PTEN， BCL2L11， and MAPK1. The roles of these key mRNAs and proteins within PI3K/Akt pathway were further validated. In comparison to the blank group， the expression levels of PTEN， BCL2L11 and MAPK1 mRNA in the hippocampus of rats in the model group were increased （P<0.01）， while the protein expression levels of p-PI3K， p-Akt and Bcl-2 were decreased （P<0.01）， and the protein expression levels of PTEN， Bax and cleaved Caspase-3 were increased （P<0.01）. Compared with the model group， the high-dose and medium-dose groups of Hei Xiaoyaosan could down-regulate the expressions of PTEN， BCL2L11 and MAPK1 mRNAs （P<0.01）， up-regulate the expressions of p-PI3K， p-Akt and Bcl-2 proteins （P<0.01）， and down-regulate the protein expressions of PTEN， Bax and cleaved Caspase-3 （P<0.05， P<0.01）. ConclusionHei Xiaoyaosan may regulate PI3K/Akt signaling pathway by down-regulating expressions of key genes such as PTEN， BCL2L11 and MAPK1， and thus improve the learning and memory abilities of insomnia rats with liver depression syndrome.

Charcoal-processed traditional Chinese herbal medicine has various therapeutic effects， including astringing， hemostasis， anti-diarrhea， clearing heat， tonifying， and warming the interior. This paper summarizes the clinical application features， compatible experiences， dosages， and precautions for over 20 types of charcoal-processed herbal medicine in the treatment of gynecological bleeding disorders caused by dysfunctions such as dysfunctional uterine bleeding， endometriosis， uterine incision pseudocavity， and vaginal bleeding resulting from threatened miscarriage. The charcoal-processed herbal medicine include Huangqin （Scutellaria Baicalensis） Charcoal， Dahuang （Rheum Palmatum） Charcoal， Cebai （Platycladus Orientalis） Charcoal， Diyu （Sanguisorba Officinalis） Charcoal， Daji （Cirsium Setosum） Charcoal， Xiaoji （Cirsium Japonicum） Charcoal， Shengdi （Rehmannia Glutinosa） Charcoal， Aiye （Artemisia Argyi） Charcoal， Paojiang （Zingiber Officinale） Charcoal， Xuduan （Dipsacus Asper） Charcoal， Duzhong （Eucommia Ulmoides） Charcoal， Qiancao （Rubia Cordifolia） Charcoal， Puhuang （Typha Angustifolia） Charcoal， Shanzha （Crataegus Pinnatifida） Charcoal， Jingjie （Schizonepeta Tenuifolia） Charcoal， Xueyu （Carthamus Tinctorius） Charcoal， Zonglyu （Areca Catechu） Charcoal， Wumei （Prunus Mume） Charcoal， Shudahuang （Rheum Officinale） Charcoal， Lianfang （Nymphaea Alba） Charcoal， Mianmaguanzhong （Clematis Armandii） Charcoal， and Oujie （Nelumbo Nucifera） Charcoal.

ObjectiveTo investigate the mechanism of action of Bushen Huoxue prescription （BSHXP） in regulating premature ovarian failure in rats through the PTEN-induced kinase 1 （PINK1）/Parkinson's protein （Parkin） signaling pathway-mediated mitophagy. MethodsA total of 48 rats were randomly divided into a blank group consisting of eight rats， while the remaining 40 rats underwent modeling. The modeling group was intraperitoneally injected with 4 mg·kg^-1 cisplatin solution， followed by a second injection one week later， for a total of two injections. The estrous cycle was observed through vaginal smears for 14 consecutive days to determine whether the modeling was successful. The successfully modeled rats were randomly divided into a model group， groups receiving low， medium， and high doses of BSHXP at 9.72， 19.44， and 38.88 g·kg^-1·d^-1 （BSHXP-L， BSHXP-M， and BSHXP-H groups）， and a positive control group treated with estradiol valerate （0.09 mg·kg^-1·d^-1）， for 21 consecutive days. The body weight of the rats was measured weekly. After the final administration， rats were anesthetized， and their blood and ovaries were collected. The ovarian weight was measured. Enzyme-linked immunosorbent assay （ELISA） was used to detect serum levels of anti-Müllerian hormone （AMH）， follicle-stimulating hormone （FSH）， luteinizing hormone （LH）， and estradiol （E₂）. Assay kits were used to measure the levels of superoxide dismutase （SOD） and malondialdehyde （MDA） in the rat serum. Hematoxylin-eosin （HE） staining was used to observe the morphological changes in the ovaries. Immunohistochemistry （IHC） was performed to detect microtubule autophagy-related protein 1 light chain 3B（LC3B） protein expression in ovarian tissue， and electron microscopy was employed to examine the mitochondrial and autophagosome changes in the rat ovaries. Western blot was used to detect the protein expression of PINK1， Parkin， LC3B， and p62. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression of PINK1， Parkin， LC3B， and p62 in ovarian tissue. ResultsCompared with the blank group， the model group showed significant reductions in body weight， weight gain， and ovarian weight （P<0.01）， along with decreased serum AMH and E₂ levels （P<0.01）， while FSH and LH levels were increased （P<0.01）. Serum MDA levels were significantly increased （P<0.01）， and SOD levels were significantly reduced （P<0.01）. The ovarian tissue structure was disordered， and the zona pellucida was wrinkled into an irregular acidophilic annular object， accompanied by an increased number of closed follicles. Electron microscopy showed mitochondrial swelling， unclear structure， and no obvious autophagosomes and autolysosome structures. The proteins and mRNA expression levels of PINK1， Parkin， LC3B， and p62 in the ovarian tissue were significantly reduced （P<0.01）. Compared with the model group， all treatment groups showed varying degrees of increases in body weight and ovarian weight （P<0.05， P<0.01）. Except for the BSHXP-L group， all treatment groups showed increased body weight gain （P<0.01）. All treatment groups showed significantly increased serum AMH and decreased FSH levels （P<0.01）. Except for the BSHXP group， all treatment groups showed varying degrees of increase and decrease in serum E₂ and LH levels （P<0.05， P<0.01）. All treatment groups showed reduced serum MDA levels （P<0.01）， while the BSHXP-M， BSHXP-H， and the positive control groups demonstrated improved serum SOD levels （P<0.05， P<0.01）. All treatment groups showed an increased number of follicles at all stages， visible mature follicles， and a decreased number of closed follicles. Electron microscopy showed relieved mitochondrial swelling， morphology close to normal， clear structure， and visible formation of autolysosomes in all treatment groups. Additionally， the protein and mRNA expression levels of PINK1， Parkin， LC3B， and p62 in ovarian tissue were significantly increased （P<0.05， P<0.01）. ConclusionBSHXP may improve ovarian function in rats with premature ovarian failure by regulating the PINK1/Parkin signaling pathway， activating mitochondrial autophagy， and reducing oxidative damage.

ObjectiveTo investigate the mechanism of action of Bushen Huoxue prescription （BSHXP） in regulating premature ovarian failure in rats through the PTEN-induced kinase 1 （PINK1）/Parkinson's protein （Parkin） signaling pathway-mediated mitophagy. MethodsA total of 48 rats were randomly divided into a blank group consisting of eight rats， while the remaining 40 rats underwent modeling. The modeling group was intraperitoneally injected with 4 mg·kg^-1 cisplatin solution， followed by a second injection one week later， for a total of two injections. The estrous cycle was observed through vaginal smears for 14 consecutive days to determine whether the modeling was successful. The successfully modeled rats were randomly divided into a model group， groups receiving low， medium， and high doses of BSHXP at 9.72， 19.44， and 38.88 g·kg^-1·d^-1 （BSHXP-L， BSHXP-M， and BSHXP-H groups）， and a positive control group treated with estradiol valerate （0.09 mg·kg^-1·d^-1）， for 21 consecutive days. The body weight of the rats was measured weekly. After the final administration， rats were anesthetized， and their blood and ovaries were collected. The ovarian weight was measured. Enzyme-linked immunosorbent assay （ELISA） was used to detect serum levels of anti-Müllerian hormone （AMH）， follicle-stimulating hormone （FSH）， luteinizing hormone （LH）， and estradiol （E₂）. Assay kits were used to measure the levels of superoxide dismutase （SOD） and malondialdehyde （MDA） in the rat serum. Hematoxylin-eosin （HE） staining was used to observe the morphological changes in the ovaries. Immunohistochemistry （IHC） was performed to detect microtubule autophagy-related protein 1 light chain 3B（LC3B） protein expression in ovarian tissue， and electron microscopy was employed to examine the mitochondrial and autophagosome changes in the rat ovaries. Western blot was used to detect the protein expression of PINK1， Parkin， LC3B， and p62. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression of PINK1， Parkin， LC3B， and p62 in ovarian tissue. ResultsCompared with the blank group， the model group showed significant reductions in body weight， weight gain， and ovarian weight （P<0.01）， along with decreased serum AMH and E₂ levels （P<0.01）， while FSH and LH levels were increased （P<0.01）. Serum MDA levels were significantly increased （P<0.01）， and SOD levels were significantly reduced （P<0.01）. The ovarian tissue structure was disordered， and the zona pellucida was wrinkled into an irregular acidophilic annular object， accompanied by an increased number of closed follicles. Electron microscopy showed mitochondrial swelling， unclear structure， and no obvious autophagosomes and autolysosome structures. The proteins and mRNA expression levels of PINK1， Parkin， LC3B， and p62 in the ovarian tissue were significantly reduced （P<0.01）. Compared with the model group， all treatment groups showed varying degrees of increases in body weight and ovarian weight （P<0.05， P<0.01）. Except for the BSHXP-L group， all treatment groups showed increased body weight gain （P<0.01）. All treatment groups showed significantly increased serum AMH and decreased FSH levels （P<0.01）. Except for the BSHXP group， all treatment groups showed varying degrees of increase and decrease in serum E₂ and LH levels （P<0.05， P<0.01）. All treatment groups showed reduced serum MDA levels （P<0.01）， while the BSHXP-M， BSHXP-H， and the positive control groups demonstrated improved serum SOD levels （P<0.05， P<0.01）. All treatment groups showed an increased number of follicles at all stages， visible mature follicles， and a decreased number of closed follicles. Electron microscopy showed relieved mitochondrial swelling， morphology close to normal， clear structure， and visible formation of autolysosomes in all treatment groups. Additionally， the protein and mRNA expression levels of PINK1， Parkin， LC3B， and p62 in ovarian tissue were significantly increased （P<0.05， P<0.01）. ConclusionBSHXP may improve ovarian function in rats with premature ovarian failure by regulating the PINK1/Parkin signaling pathway， activating mitochondrial autophagy， and reducing oxidative damage.

ObjectiveBased on ultra-high performance liquid chromatography-quadrupole-time-of-flight mass spectrometry（UPLC-Q-TOF-MS）， network pharmacology and pharmacodynamics， to investigate the pharmacodynamic material basis and mechanism of Hei Xiaoyaosan in improving learning and memory ability of insomnia rats. MethodUPLC-Q-TOF-MS was used to characterize the chemical constituents of Hei Xiaoyaosan. Network pharmacology was applied to construct the network of active ingredients-intersecting targets-pathways， and molecular docking was performed on key ingredients and core targets. Sixty 8-week-old male SD rats were selected and randomly divided into blank group， model group， Hei Xiaoyaosan low， medium， and high dose groups（3.82， 7.65， 15.30 g·kg^-1）， and zolpidem tartrate group（0.5 mg·kg^-1）， with 10 rats in each group. Except for the blank group， the insomnia model was induced by intraperitoneal injection of p-chlorophenylalanine（PCPA） for 4 consecutive days. Rats in each dosing group were administered the corresponding dose by gavage， once a day for 14 consecutive days. Morris water maze test was utilized to assess the learning and memory ability of rats， transmission electron microscopy was employed to examine the ultrastructure of hippocampal synapses， enzyme-linked immunosorbent assay（ELISA） was conducted to analyze the levels of 5-hydroxytryptamine（5-HT）， interleukin-6（IL-6）， and tumor necrosis factor-α（TNF-α） in hippocampal tissues， and Western blot was performed to detect the expression levels of tumor suppressor protein p53（TP53）， rat sarcoma virus（RAS）， epidermal growth factor receptor（EGFR）， cyclic adenosine monophosphate（cAMP）-response element binding protein（CREB） binding protein（CREBBP）， glycogen synthase kinase-3β（GSK-3β）， protein kinase B1（Akt1）， nitric oxide synthase 1（NOS1）， phosphorylated（p）-Akt1， and p-GSK-3β in hippocampal tissues. Additionally， real-time fluorescence quantitative polymerase chain reaction（Real-time PCR） was used to assess the mRNA expression levels of TP53， RAS， EGFR， CREBBP， GSK-3β， Akt1 and NOS1. ResultA total of 176 components were identified in Hei Xiaoyaosan， mainly flavonoids， triterpene saponins， phenylpropanoids and other compounds. Network pharmacological analysis revealed that TP53， V-Ha-Ras Harvey Rat sarcoma viral oncogene homolog（HRAS）， neuroblastoma sarcoma viral oncogene homolog（NRAS）， EGFR， CREBBP， GSK-3β， Akt1 and NOS1 were the key targets of Hei Xiaoyaosan in treating insomnia. The core targets were predominantly associated with cAMP， RAS， Ras-associated protein 1（Rap1）， advanced glycation end products（AGE）/receptor for AGE（RAGE）， and EGFR signaling pathways， and the key active ingredients of Hei Xiaoyaosan in treating insomnia were 8-shogaol， ligustilide F， 6-gingerol， levistilide A and senkyunolide E. Animal experiment results demonstrated that Hei Xiaoyaosan medium and high dose groups significantly increased body weight， shortened sleep latency and prolonged sleep duration in insomnia rats（P<0.01）， significantly decreased escape latency and increased platform crossing frequency（P<0.01）， and improved the pathological changes of hippocampal synaptic ultrastructure. Meanwhile， the two groups could significantly elevate 5-HT level， Akt1 mRNA expression， Akt1 and p-Akt1 protein expression（P<0.01）， reduce inflammatory factor levels（P<0.01）， and down-regulate protein expression levels of TP53， RAS， NOS1， EGFR， CREBBP， GSK-3β and p-GSK-3β（P<0.01）， as well as mRNA expression levels of TP53， RAS， NOS1， EGFR， CREBBP and GSK-3β in hippocampal tissues（P<0.01）. ConclusionThis study determined that the five key active ingredients（8-shogaol， ligustilide F， 6-gingerol， levistilide A and senkyunolide E） in Hei Xiaoyaosan may improve the learning and memory ability of insomnia rats by regulating signaling pathways such as cAMP， RAS， and EGFR， providing an important reference for its mechanism research and clinical application.

This article summarized Professor You Zhaoling's clinical experience in the treatment of polycystic ovary syndrome(PCOS).It is believed that kidney deficiency and blood stasis are the basic etiology and pathogenesis,with liver and spleen disorders.Clinical treatment is based on tonifying the kidney and replenishing essence,promoting blood circulation and resolving stasis,supplemented by soothing the liver,strengthening the spleen,and resolving dampness;treatment should also be based on the patients'stage of illness,age,and body type;basic body temperature and B-ultrasound should be skillfully uses to monitor ovulation,and four methods of regulating bubbles were creatively created;an original hypothetical menstrual stimulation method for PCOS patients with menstrual disorders was created.Professor You's clinical experience has demonstrated the characteristics of TCM treatment,providing a new approach for the TCM treatment of PCOS.

Existing antidepressant treatments are generally suboptimal for patients with bipolar disorder(BD).Several studies have explored the efficacy of light therapy(LT)in patients with bipolar depression,along with investigating parameters,devices,and safety aspects of LT.This paper provides a review of these aspects.Numerous meta-analyses and randomized controlled trials have indicated that LT could significantly improve depressive symptoms in BD patients,with both low and high intensity white light having this effect,while the antidepressant effect of blue light remains unclear.LT takes effect rapidly,preferably in the morning,with each session lasting between 30 to 60 minutes,but there is no consensus on the most beneficial LT course for BD patients.The most commonly used device for LT is a lightbox.However further exploration is need regarding the safety of LT glasses.When LT devices that meet safety standards is selected,the overall safety of LT will be high and risk of manic or hypomanic switch will be low for BD patients.In conclusion,LT holds promise for patients with bipolar depression,and further research on LT for BD patients should be conducted to explore LT strategies and develop LT prescriptions.

Objective To investigate the expression levels of plasma circular RNA PTP4A2(circPTP4A2)and circTLK2 in elderly patients with acute ischemic stroke(AIS)and their predic-tive value for neurological functional outcomes.Methods A total of 122 elderly AIS patients admitted to our department from May 2021 to December 2022 were prospectively recruited,and according to their modified Rankin Scale(mRS)score at 3 months after stroke onset,they were divided into a good outcome group(mRS score≤2,81 cases)and a poor outcome group(mRS score:3-6,41 cases).Their baseline data,and plasma circPTP4A2 and circTLK2 levels were compared between the two groups.Multivariate logistic regression analysis was employed to iden-tify prognostic factors for poor outcomes in the elderly AIS patients.ROC curve analysis was ap-plied to evaluate the prognostic value of circPTP4A2 and circTLK2 for adverse outcomes in the patients.Pearson correlation analysis was performed to assess the relationship of plasma levels of circPTP4A2 and circTLK2 with NIHSS score,as well as mRS score.Results The plasma expres-sion levels of circPTP4A2 and circTLK2 were significantly higher in the poor outcome group than the good outcome group[2.08(0.87,2.77)vs 0.93(0.63,1.20),1.71(0.92,2.80)vs 0.75(0.49,1.09),P＜0.01].Multifactor logistic regression analysis showed that plasma circPTP4A2 and circTLK2 were independent predictive factors for poor functional outcomes in elderly AIS patients(P＜0.01,P＜0.05).ROC curve analysis demonstrated that the AUC value of combined circPTP4A2 and circTLK2 in predicting poor outcome in elderly AIS patients was 0.787(95％CI:0.691-0.883).Pearson correlation analysis revealed that the expression levels of circPTP4A2 and cir-cTLK2 in elderly AIS patients were mildly positively correlated with baseline NIHSS scores(r=0.463,r=0.456;P＜0.01)and moderately positively correlated with mRS scores at 3 months after stroke onset(r=0.682,r=0.604;P＜0.01).Conclusion Plasma circPTP4A2 and circTLK2 may be potential biomarkers for predicting neurological functional outcomes in elderly AIS patients.

BACKGROUND: Currently, a significant number of miners are involved in mining operations at the Gejiu tin mine in Yunnan. This occupational setting is associated with exposure to dust particles, heavy metals, polycyclic aromatic hydrocarbons, and radioactive radon, thereby significantly elevating the risk of lung cancer. This study aims to investigate the involvement of leptin-mediated extracellular regulated protein kinase (ERK) signaling pathway in the malignant transformation of rat alveolar type II epithelial cells induced by Yunnan tin mine dust. METHODS: Immortalized rat alveolar cells type II (RLE-6TN) cells were infected with Yunnan tin mine dust at a concentration of 200 μg/mL for nine consecutive generations to establish the infected cell model, which was named R₂₀₀ cells. The cells were cultured normally, named as R cells. The expression of leptin receptor in both cell groups was detected using the Western blot method. The optimal concentration of leptin and mitogen-activated protein kinase kinase (MEK) inhibitor (U0126) on R₂₀₀ cells was determined using the MTT method. Starting from the 20th generation, the cells in the R group were co-cultured with leptin, while the cells in the R₂₀₀ group were co-cultured with the MEK inhibitor U0126. The morphological alterations of the cells in each group were visualized utilizing hematoxylin-eosin staining. Additionally, concanavalin A (ConA) was utilized to detect any morphological differences, and an anchorage-independent growth assay was conducted to assess the malignant transformation of the cells. The changes in the ERK signaling pathway in epithelial cells after the action of leptin were detected using the Western blot method. RESULTS: Both the cells in the R group and R₂₀₀ group express leptin receptor OB-R. Compared to the R₂₀₀ group, the concentration of leptin at 100 ng/mL shows the most significant pro-proliferation effect. The proliferation of R₂₀₀ cells infected with the virus is inhibited by 30 μmol/L U0126, and a statistically significant divergence was seen when compared to the control group (P<0.05). Starting from the 25th generation, the cell morphology of the leptin-induced R₂₀₀ group (R₂₀₀L group) underwent changes, leading to malignant transformation observed at the 30th generation. The characteristics of malignant transformation became evident by the 40th generation in the R₂₀₀L group. In contrast, the other groups showed agglutination of P40 cells, and the speed of cell aggregation increased with an increase in ConA concentration. Notably, the R₂₀₀L group exhibited faster cell aggregation compared to the U0126-induced R₂₀₀ (R₂₀₀LU) group. Additionally, the cells in the R₂₀₀L group were capable of forming clones starting from P30, with a colony formation rate of 2.25‰±0.5‰. However, no clonal colonies were observed in the R₂₀₀LU group and R₂₀₀ group. The expression of phosphorylated extracellular signal-regulated kinase (pERK) was enhanced in cells of the R₂₀₀L group. However, when the cells in the R₂₀₀L group were treated with U0126, a blocking agent, the phosphorylation level of pERK decreased. CONCLUSIONS Leptin can promote the malignant transformation of lung epithelial cells infected by mine dust, and the ERK signaling pathway may be necessary for the transformation of alveolar type II epithelial cells induced by Yunnan tin mine dust.
Rats ; Animals ; Alveolar Epithelial Cells/pathology* ; Dust ; Tin/adverse effects* ; Lung Neoplasms/pathology* ; Leptin/adverse effects* ; Receptors, Leptin ; China ; Signal Transduction ; Epithelial Cells/pathology* ; Mitogen-Activated Protein Kinase Kinases/adverse effects*

Objective To investigate the role of chemokine ligand 19 (CCL19) and protein kinase-B (AKT) signaling pathway in lung cancer development. Methods The human lung adenocarcinoma cell line, A549 cells, in logarithmic growth phase were randomly divided into five groups： blank control group, solvent control group, CCL19 treatment group, AKT inhibition group, and antibody neutralization group. The blank control group received no treatment. The other four groups were treated with dimethyl sulfoxide, CCL19, MK-2206 (AKT inhibitor), and a combination of CCL19 and MK-2206, respectively. Cell viability was assessed using the CCK-8 assay, while cell migration and invasion capabilities were evaluated using the cell scratch and transwell assays. The relative expression levels of Pan-AKT, p-AKT (Ser473), p-AKT (Thr308), E-cadherin (E-cad), N-cadherin (N-cad), and Snail proteins in A549 cells were detected using Western blotting. Lung cancer tissue samples from 60 patients with non-small cell lung cancer (NSCLC) were collected, and the expression of CCL19 and matrix metalloproteinase 9 (MMP9) proteins in the specimens was examined using immunohistochemistry. Results The survival rate of A549 cells in the AKT inhibition group and antibody neutralization group was lower than that in blank control group, solvent control group, and CCL19 treatment group (all P<0.05). The cell scratch assay result showed that the cell migration rate of the CCL19 treatment group was higher at 36.0 and 48.0 hours than those of the blank control group, solvent control group, AKT inhibition group, and neutralizing antibody group (all P<0.05). The Transwell assay result showed that the invasion amount of A549 cells in the AKT inhibition group was less than that in the CCL19 treatment group (P<0.05). Compared with the blank control group, the relative expression of E-cad protein in the CCL19 treatment group decreased, while the relative expression of p-AKT (Ser473), p-AKT (Thr308), N-cad and Snail proteins increased (all P<0.05). The relative expression of p-AKT (Ser473), p-AKT (Thr308), N-cad, and Snail proteins in A549 cells decreased (all P<0.05), and relative expression of E-cad protein increased (all P<0.05) in the AKT inhibition group and antibody neutralization group compared with the blank control group, solvent control group, and CCL19 treatment group. There was no significant difference in the expression of CCL19 and MMP9 in lung cancer tissues of NSCLC patients in Xuanwei City, Gejiu City, and other regions (all P>0.05). The expression of CCL19 and MMP9 in NSCLC patients with lymph node metastasis was higher than in patients without lymph node metastasis (all P<0.01). Conclusion CCL19 can promote the invasion and metastasis of lung cancer cells and induce epithelial-mesenchymal transition. Its expression level is related to lymph node metastasis in NSCLC patients. The AKT signaling pathway may be an important mechanism underlying lung cancer development.