ObjectiveTo study the effect and mechanism of fibrinogen （Fib） overexpression on mitochondrial quality control system in the rat model of coronary heart disease with the syndrome of blood stasis. MethodsForty male SD rats were randomly assigned into normal， model， Fib， and empty vector （AAV） groups， with 10 rats in each group. The model， Fib， and AAV groups were fed with a high-fat diet adaptively and administrated with 3×10⁶ U·kg^-1 vitamin D3 powder by gavage after 7 days and 2×10⁶ U·kg^-1 vitamin D3 solution after 14 days. After being fed with a high-fat diet for 7 weeks， rats in each group received subcutaneous injection of isoproterenol （5 mg·kg^-1） for 3 days. During the modeling period， rats in the normal group were fed with ordinary feed without any special treatment. The changes in blood lipid and hemorheological indexes of rats in each group were measured. The aorta tissue was stained with hematoxylin-eosin （HE）， and the standard lead Ⅱ electrocardiograms （ECGs） of rats in each group were recorded. Enzyme-linked immunosorbent assay （ELISA） and real-time PCR were employed to verify the overexpression levels of Fib in the liver and plasma. Western blotting was employed to determine the protein levels of mitofusin 2 （Mfn2）， optic atrophy protein 1 （OPA1）， dynamin-related protein 1 （Drp1）， phosphorylated adenosine monophosphate-activated protein kinase （p-AMPK）/adenosine monophosphate-activated protein kinase （AMPK）， peroxisome proliferator-activated receptor γ-coactivator-1α （PGC-1α）， PTEN-induced putative kinase 1， and Parkin. Real-time PCR was employed to determine the mRNA levels of AMPK and PGC-1α in the myocardial tissue. The changes in levels of adenosine triphosphate （ATP） and adenosine monophosphate （AMP） in the myocardial tissue were determined by ELISA. ResultsCompared with the normal group， the other three groups showed elevated levels of total cholesterol and low-density lipoprotein cholesterol （P<0.01） and no significant changes in levels of triglyceride and high-density lipoprotein cholesterol. Compared with the model group， the Fib and AAV groups showed risen levels of total cholesterol （P<0.05， P<0.01）. Compared with the normal group， the model and Fib groups presented increases in low shear viscosity and middle shear viscosity （P<0.05， P<0.01）， and the Fib group showcased an increase in high shear viscosity （P<0.01）. Compared with the model group， the Fib group showed increases in low shear viscosity， middle shear viscosity， and high shear viscosity （P<0.05， P<0.01）. Compared with the Fib group， the AAV group demonstrated decreases in low shear viscosity， middle shear viscosity， and high shear viscosity （P<0.05， P<0.01）. The normal group had an complete aortic structure with well arrangement of elastic fibers. In the model group， the vascular wall became thickened and the intima was rough with inflammatory infiltration. In the Fib group， the intima calcification formed a cavity structure and the intima was abnormally proliferated， while in the AAV group， the intima smooth muscle was slightly proliferated with local calcification. The ECG of the normal group indicated sinus rhythm， and that of the model group presented ST segment oblique elevation （>0.1 mV）. The ECG of the Fib group presented characteristic ST segment arch back elevation with T-wave towering， and that of the AAV group presented ST segment oblique elevation. Compared with the normal group， the model and Fib groups showed elevations in levels of liver Fib， plasma Fib， and liver Fibα mRNA （P<0.01）， and the AAV group had risen levels of Fib and Fibα mRNA （P<0.01）. Compared with the model group， the Fib group presented risen levels of liver Fib and Fibα mRNA （P<0.01）. Compared with the Fib group， the AAV group presented decreases in levels of liver Fib， plasma Fib， and liver Fibα mRNA （P<0.01）. Compared with the normal group， the other three groups had down-regulated protein and mRNA levels of Mfn2， OPA1， PINK1， Parkin， p-AMPK/AMPK， and PGC-1α （P<0.05， P<0.01） and up-regulated protein levels of Drp1 （P<0.01）. Compared with those in the model group， the mRNA and protein levels of Mfn2， OPA1， PINK1， Parkin， p-AMPK/AMPK， and PGC-1α were all down-regulated （P<0.05， P<0.01） and the protein level of Drp1 was up-regulated （P<0.01） in the Fib group. Compared with the Fib group， the AAV group showed differences in protein levels of OPA1， PGC-1α， Parkin， and Drp1 （P<0.05， P<0.01） and an increasing trend in the mRNA levels of AMPK and PGC-1α with no significant difference. Compared with the normal group， the other three groups had elevated levels of ATP in the myocardial tissue （P<0.01）. Compared with the model group， the Fib group showed elevated levels of ATP and AMP （P<0.01）. Compared with the Fib group， the AAV group exhibited lowered levels of ATP and AMP （P<0.01）. ConclusionFib can achieve the overexpression effect in the rat model of coronary heart disease with the syndrome of blood stasis. At the same time， the overexpression of Fib can induce the damage of the mitochondrial quality control system in the myocardial tissue， inhibit mitochondrial dynamics and mitochondrial biosynthesis， and down-regulate mitochondrial autophagy， thereby aggravating myocardial injury in the rat model.

ObjectiveTo analyze the metabolomic characteristics of platelets in fibrinogen（FIB） overexpression rats of coronary heart disease with blood stasis syndrome（CHD-BSS）， explore potential biomarkers， and investigate the mechanism of FIB overexpression on CHD-BSS. MethodsSD rats were randomly divided into BSS group and BSS+FIB overexpression group（BSS+FIB group）， with 10 rats in each group. Both the BSS+FIB group and the BSS group were fed a high-fat diet combined with oral administration of vitamin D₃ and subcutaneous injection of isoproterenol， but rats in the BSS+FIB group were overexpressed with FIB during the initial modeling stage by transfection with adeno-associated virus（AAV）. The overexpression level of FIB in rat liver and plasma samples was detected by enzyme-linked immunosorbent assay（ELISA） and real-time fluorescence quantitative polymerase chain reaction（Real time PCR）， as well as the expression level of liver FIB A（FGA） mRNA. The characteristics of metabolites in rat platelet samples were analyzed by ultra-high performance liquid chromatography-quadrupole-electrostatic field orbital trap high-resolution mass spectrometry（UPLC-Q-Exactive Orbitrap-MS）， and the differential metabolites between groups were screened by principal component analysis（PCA） and orthogonal partial least squares-discriminant analysis（OPLS-DA）， and the enriched pathways were analyzed. The accuracy of potential biomarkers in the diagnosis of CHD-BSS was evaluated by receiver operating characteristic（ROC） curve. The expression of autophagy related proteins phosphorylated adenosine monophosphate（AMP） activated protein kinase（p-AMPK）/AMPK， phosphorylated mammalian target of rapamycin（p-mTOR）/mTOR， microtubule-associated protein 1 light chain 3（LC3） Ⅱ/Ⅰ and p62 in platelets were detected by Western blot. ResultsCompared with the BSS group， the expression levels of FIB in liver and plasma samples of the BSS+FIB group were significantly increased（P<0.05， P<0.01）， and the expression level of FIB mRNA in the liver was remarkably increased（P<0.01）， indicating successful overexpression of FIB. Platelet metabolomics results showed significant differences in metabolic profiles between the BSS+FIB group and the BSS group， and a total of 25 significantly enriched metabolic pathways and 8 metabolites involved in these metabolic pathways， among which uric acid， guanosine and ribose 1-phosphate levels were up-regulated， while adenosine diphosphate（ADP）， AMP， guanosine diphosphate（GDP）， adenylosuccinate and norepinephrine levels were down-regulated. The diagnostic ability analysis of differential metabolites showed that all 8 differential metabolites had good diagnostic ability， with an area under the curve（AUC）>0.85. Western blot results showed that compared with the BSS group， the expression levels of p-mTOR/mTOR and p62 proteins in platelets of the BSS+FIB group was significantly reduced（P<0.01）， while the expression levels of p-AMPK/AMPK and LC3Ⅱ/Ⅰ proteins were increased， but the difference was not statistically significant. ConclusionOverexpression of FIB can change the metabolic characteristics of CHD-BSS rat model， involving multiple aspects such as vascular endothelial injury， platelet activation and myocardial function damage. Among them， overexpression of FIB may enhance the occurrence of platelet autophagy， thereby inducing platelet activation and promoting thrombus formation.

Objective:To carry out carrier screening among people of childbearing age, detect the pathogenic genes of monogenic genetic diseases and analyze the carrier status of pathogenic variants, so as to provide fertility guidance and intervention measures for high-risk families.Methods:From August 2022 to August 2023, 1 533 families of childbearing age who met the criteria were recruited in the Chinese PLA General Hospital, including a total of 3 044 subjects. According to the standard enrollment procedure, 223 genes (197 autosomal recessive genes and 26 X-linked genes) of the subjects were tested. According to the screening results, genetic counseling and fertility guidance were provided to the subjects. Invasive prenatal diagnosis was performed for high-risk couples (both couples being carriers of the same autosomal recessive disease gene or the woman was a carrier of X-linked disease gene), and their pregnancy pattern, outcome and offspring phenotype were followed up.Results:(1) A total of 3 044 cases from 1 511 couples and women of childbearing age from 22 families were included for carrier screening. Totally 1 503 families chose simultaneous screening and 30 families chose sequential screening out of the 1 533 families. Among the 3 044 subjects, 1 603 individuals carried at least one pathogenic or likely pathogenic variant, and the overall carrier rate was 52.66% (1 603/3 044). A total of 2 292 pathogenic or likely pathogenic variants were detected, and 0.75 variants (2 292/3 044) were detected per capita. (2) The three genes with the highest carrier rates were GJB2 (8.67%, 264/3 044), CYP21A2 (3.19%, 97/3 044) and PAH (3.09%, 94/3 044). There were 32 genes with a carrier rate ≥1/200, 17 genes with a carrier rate ≥1/100, and 7 genes with a carrier rate ≥1/50. (3) Thirty-eight high-risk families were identified. After excluding G6PD gene mutation, there were 33 high-risk families, of which 25 couples were carriers of the same autosomal recessive gene, 9 women were carriers of X-linked gene, and 1 family was double high-risk couple with both autosomal recessive and X-linked gene. After further excluding the GJB2 c.109G>A mutation, 21 high-risk families were identified. Preimplantation genetic testing for monogenic disease was performed in 12 families after genetic counseling. Prenatal diagnosis was completed in 4 out of 5 high-risk families who conceived naturally. Two fetuses carried the parental variants and terminated the pregnancy, one fetus did not carry the parental variants but was induced due to trisomy 21 syndrome, and one fetus was a carrier of congenital disorders of glycosylation type 1a.Conclusions:Carrier screening effectively identifies high-risk genetic disease families and provides reproductive guidance to prevent the birth of affected children. However, establishing multidisciplinary team is essential for managing complex cases. Implementation should prioritize prenatal institutions with genetic counseling or diagnostic expertise for monogenic disorders or established referral networks.

Objective:To investigate the effect of tumor necrosis factor α (TNF-α) on the permeability of blood labyrinth barrier in C57BL/6J male mice and its possible mechanism.Methods:As for the design of animal experiment, twenty male C57BL/6J mice aged 6-8 weeks were randomly divided into control group and cisplatin group with 10 mice in each group by software method. The control group was intraperitoneally injected with normal saline every day, and the cisplatin group was intraperitoneally injected with 4 mg/kg cisplatin for 4 consecutive days. After administration, auditory brainstem response (ABR) was used to detect hearing changes in mice. HE staining was used to observe the morphological changes of mouse cochlea vasculature. The expression of TNF-α was detected by immunohistochemistry and ELISA. The permeability of the blood labyrinth barrier was observed by Evans blue staining. With respect to the design of cell experiment, primarily cultured cochlea pericytes (PC) and endothelial cells (EC) were divided into EC group, EC+TNF-α group, EC+PC group, EC+PC+TNF-α group, EC+PC+TNF-α+SB-3CT (MMP-9/MMP-2 secretion inhibitor) group, PC group, PC+TNF-α group, PC+TNF-α+LY294002 (PI3K/AKT pathway inhibitor) group, PC+LY294002 group. The protein expressions of ZO-1, VE-cadherin, MMP-9, MMP-2, PI3K, p-PI3K, AKT, and p-AKT were detected by Western blot. TEER and FITC-dextran penetration experiment were used to detect EC resistance and monolayer EC permeability, respectively. The data were statistically analyzed using GraphPad Prism 8 software.Results:In animal experiment, compared with control group, the ABR response threshold of mice in cisplatin group was increased ( P<0.01). The vaccine ular structure of the cochlea was disordered red, wrinkled and vacuole increased. The extravasation of vascular red fluorescent dye increased ( P<0.05), and also, levels of serum TNF-α and cochlear veins increased ( P<0.01). In cell experiment, by treatment of EC with different concentrations of cisplatin, 20 μmol cisplatin led to the highest expression of TNF-α ( P<0.01). The expression of TNF-α was the highest after 3 h intervention in EC ( P<0.05). Compared with those in EC+PC group, the resistance value of EC was decreased, the permeability of monolayers EC was increased, the expression of ZO 1 and VE cadherin proteins was decreased, and however, the resistance value was increased and the permeability of EC was decreased after the intervention of SB-3CT in EC+PC+TNF-α group. The expressions of ZO-1 and VE-cadherin proteins were increased ( P<0.05). The expression of MMP-9 increased after TNF-α intervention ( P<0.05), the expression of MMP-2 had no significant change, and the expression of p-PI3K/PI3K and p-AKT/AKT were increased ( P<0.05). The expression of MMP-9 decreased in PC+TNF-α+LY294002 group ( P<0.05). Conclusion:The hearing loss of C57BL/6J male mice induced by cisplatin may be related to the increased release of TNF-α from the blood labyrinth barrier EC in the cochlear stria vascularis, and the activation of PI3K/AKT signaling pathway by TNF-α in PC, which increases the secretion of MMP-9 from PC, ultimately leads to the increased permeability of the blood labyrinth barrier.

Objective:To carry out carrier screening among people of childbearing age, detect the pathogenic genes of monogenic genetic diseases and analyze the carrier status of pathogenic variants, so as to provide fertility guidance and intervention measures for high-risk families.Methods:From August 2022 to August 2023, 1 533 families of childbearing age who met the criteria were recruited in the Chinese PLA General Hospital, including a total of 3 044 subjects. According to the standard enrollment procedure, 223 genes (197 autosomal recessive genes and 26 X-linked genes) of the subjects were tested. According to the screening results, genetic counseling and fertility guidance were provided to the subjects. Invasive prenatal diagnosis was performed for high-risk couples (both couples being carriers of the same autosomal recessive disease gene or the woman was a carrier of X-linked disease gene), and their pregnancy pattern, outcome and offspring phenotype were followed up.Results:(1) A total of 3 044 cases from 1 511 couples and women of childbearing age from 22 families were included for carrier screening. Totally 1 503 families chose simultaneous screening and 30 families chose sequential screening out of the 1 533 families. Among the 3 044 subjects, 1 603 individuals carried at least one pathogenic or likely pathogenic variant, and the overall carrier rate was 52.66% (1 603/3 044). A total of 2 292 pathogenic or likely pathogenic variants were detected, and 0.75 variants (2 292/3 044) were detected per capita. (2) The three genes with the highest carrier rates were GJB2 (8.67%, 264/3 044), CYP21A2 (3.19%, 97/3 044) and PAH (3.09%, 94/3 044). There were 32 genes with a carrier rate ≥1/200, 17 genes with a carrier rate ≥1/100, and 7 genes with a carrier rate ≥1/50. (3) Thirty-eight high-risk families were identified. After excluding G6PD gene mutation, there were 33 high-risk families, of which 25 couples were carriers of the same autosomal recessive gene, 9 women were carriers of X-linked gene, and 1 family was double high-risk couple with both autosomal recessive and X-linked gene. After further excluding the GJB2 c.109G>A mutation, 21 high-risk families were identified. Preimplantation genetic testing for monogenic disease was performed in 12 families after genetic counseling. Prenatal diagnosis was completed in 4 out of 5 high-risk families who conceived naturally. Two fetuses carried the parental variants and terminated the pregnancy, one fetus did not carry the parental variants but was induced due to trisomy 21 syndrome, and one fetus was a carrier of congenital disorders of glycosylation type 1a.Conclusions:Carrier screening effectively identifies high-risk genetic disease families and provides reproductive guidance to prevent the birth of affected children. However, establishing multidisciplinary team is essential for managing complex cases. Implementation should prioritize prenatal institutions with genetic counseling or diagnostic expertise for monogenic disorders or established referral networks.

Objective:To investigate the effect of tumor necrosis factor α (TNF-α) on the permeability of blood labyrinth barrier in C57BL/6J male mice and its possible mechanism.Methods:As for the design of animal experiment, twenty male C57BL/6J mice aged 6-8 weeks were randomly divided into control group and cisplatin group with 10 mice in each group by software method. The control group was intraperitoneally injected with normal saline every day, and the cisplatin group was intraperitoneally injected with 4 mg/kg cisplatin for 4 consecutive days. After administration, auditory brainstem response (ABR) was used to detect hearing changes in mice. HE staining was used to observe the morphological changes of mouse cochlea vasculature. The expression of TNF-α was detected by immunohistochemistry and ELISA. The permeability of the blood labyrinth barrier was observed by Evans blue staining. With respect to the design of cell experiment, primarily cultured cochlea pericytes (PC) and endothelial cells (EC) were divided into EC group, EC+TNF-α group, EC+PC group, EC+PC+TNF-α group, EC+PC+TNF-α+SB-3CT (MMP-9/MMP-2 secretion inhibitor) group, PC group, PC+TNF-α group, PC+TNF-α+LY294002 (PI3K/AKT pathway inhibitor) group, PC+LY294002 group. The protein expressions of ZO-1, VE-cadherin, MMP-9, MMP-2, PI3K, p-PI3K, AKT, and p-AKT were detected by Western blot. TEER and FITC-dextran penetration experiment were used to detect EC resistance and monolayer EC permeability, respectively. The data were statistically analyzed using GraphPad Prism 8 software.Results:In animal experiment, compared with control group, the ABR response threshold of mice in cisplatin group was increased ( P<0.01). The vaccine ular structure of the cochlea was disordered red, wrinkled and vacuole increased. The extravasation of vascular red fluorescent dye increased ( P<0.05), and also, levels of serum TNF-α and cochlear veins increased ( P<0.01). In cell experiment, by treatment of EC with different concentrations of cisplatin, 20 μmol cisplatin led to the highest expression of TNF-α ( P<0.01). The expression of TNF-α was the highest after 3 h intervention in EC ( P<0.05). Compared with those in EC+PC group, the resistance value of EC was decreased, the permeability of monolayers EC was increased, the expression of ZO 1 and VE cadherin proteins was decreased, and however, the resistance value was increased and the permeability of EC was decreased after the intervention of SB-3CT in EC+PC+TNF-α group. The expressions of ZO-1 and VE-cadherin proteins were increased ( P<0.05). The expression of MMP-9 increased after TNF-α intervention ( P<0.05), the expression of MMP-2 had no significant change, and the expression of p-PI3K/PI3K and p-AKT/AKT were increased ( P<0.05). The expression of MMP-9 decreased in PC+TNF-α+LY294002 group ( P<0.05). Conclusion:The hearing loss of C57BL/6J male mice induced by cisplatin may be related to the increased release of TNF-α from the blood labyrinth barrier EC in the cochlear stria vascularis, and the activation of PI3K/AKT signaling pathway by TNF-α in PC, which increases the secretion of MMP-9 from PC, ultimately leads to the increased permeability of the blood labyrinth barrier.

Repetitive transcranial magnetic stimulation (rTMS) has become an essential method in psychiatric disorders. However, many problems occurred in clinical application. This article interpreted the Association Standard T/CMEAS 011-2023'Operating Specifications for Repetitive Transcranial Magnetic Stimulation in Clinical Applications on Psychiatric Disorders′ released by the Chinese Medicine Education Association. The main content included a range of applications, normative references, terms and definitions, site specifications, equipment specifications, ability specifications of rTMS operators and rTMS process specifications.This article provided suggestions for clinical applications of rTMS on psychiatric disorders.

Repetitive transcranial magnetic stimulation (rTMS) has become an essential method in psychiatric disorders. However, many problems occurred in clinical application. This article interpreted the Association Standard T/CMEAS 011-2023'Operating Specifications for Repetitive Transcranial Magnetic Stimulation in Clinical Applications on Psychiatric Disorders′ released by the Chinese Medicine Education Association. The main content included a range of applications, normative references, terms and definitions, site specifications, equipment specifications, ability specifications of rTMS operators and rTMS process specifications.This article provided suggestions for clinical applications of rTMS on psychiatric disorders.

Autophagy， a mechanism of cell self-protection and self-renewal， is associated with the occurrence and development of lung cancer. Favorable autophagy can slow down the progression of lung cancer， while unfavorable autophagy can promote the progression. Therefore， regulating the level of autophagy is of great significance in the treatment of lung cancer. Healthy Qi deficiency and pathogenic Qi stagnation is an extension of the theory of deficiency and Qi stagnation proposed by the Academician WANG Yongyan. It refers to the pathological process that the abnormal body fluid metabolism caused by Qi deficiency of lung， spleen， and kidney results in phlegm and blood stasis. Lung cancer has the root cause of Qi deficiency of lung， spleen， and kidney and the syndrome of phlegm and blood stasis. The autophagy in lung cancer is interconnected with healthy Qi deficiency and pathogenic Qi stagnation. The Qi deficiency of lung， spleen， and kidney is the key factor for the weakening of favorable autophagy in lung cancer， which inhibits the apoptosis of tumor cells and leads to the accumulation of harmful substances. Phlegm and blood stasis is a direct factor enhancing the unfavorable autophagy in lung cancer， which promotes the autophagic death of normal cells， weakens the immunosuppressive effect of immune cells on tumor cells， and leads to the proliferation and migration of tumor cells. The combination of healthy Qi deficiency and pathogenic Qi stagnation results in the development of autophagy in an unfavorable direction and finally leads to the continuous progression of lung cancer. Therefore， the traditional Chinese medicine （TCM） treatment of lung cancer should follow the principle of reinforcing healthy Qi and expelling pathogenic Qi， removing phlegm and resolving stasis， so as to enhance favorable autophagy while inhibiting unfavorable autophagy. Such therapy can inhibit the proliferation and migration of tumor cells and promote the remission of lung cancer. According to the existing literature， Chinese medicine monomers are mainly used to treat lung cancer by regulating autophagy. The Chinese medicine intervention of autophagy in lung cancer mainly aims to promote the activation of autophagy. This may be because the favorable autophagy weakening caused by the Qi deficiency of lung， spleen， and kidney is the fundamental reason for the development of lung cancer.

More and more evidence shows that there is a close relationship between the inflammatory state and coronary heart disease. Inflammatory state triggers the damage of vascular endothelium in the early stage of coronary heart disease and ultimately mediates the formation of atherosclerotic plaque. The mechanism of occurrence and development of heart disease is of great significance. Phlegm is a pathological product formed by the subtle imbalance of the spleen and stomach in the transportation and transformation of water and grain. It is the general summary of a series of abnormally accumulated inflammatory substances， such as low density lipoprotein， inflammatory cells， and inflammatory factors. The nature of Phlegm determines the invasiveness and turbidity of Phlegm. Phlegm invades the meridians， causing damage to the meridians and gradually accumulating， which eventually causes the local meridian damage to aggravate. This process is similar to the persistent damage of the vascular endothelium caused by inflammation. Phlegm blocks the meridians， affects the operation of Qi and blood， causes Qi stagnation and blood stasis， and finally forms the outcome of heart and blood stasis. This process is similar to the mechanism of atherosclerotic plaques formed by continuous inflammatory damage. Heart blood stasis， depression and heat， heat toxin endogenous， forming the syndrome of heat toxin stasis， which is similar to the process of atherosclerotic plaque rupture and thrombosis causing acute cardiovascular events.The formation of Phlegm is rooted in the deficiency of spleen. Based on the ''phlegm，stasis，toxin'' theory， spleen deficiency is the intrinsic pathogenesis of the inflammatory state of coronary heart disease， and the invasion of phlegm， blood stasis of heart， heat and blood stasis are the evolution of inflammatory damage of coronary heart disease. Traditional Chinese medicine differentiation and treatment is based on strengthening the spleen and nourishing Qi to treat the root and removing phlegm and blood stasis， and clearing heat and detoxifying to treat symptoms. The related Chinese medicine compounds， Chinese patent medicines， and single Chinese medicines can reduce the inflammatory indicators of coronary heart disease， thereby improving the prognosis of coronary heart disease.