1.Mechanism of Yishen Huoxue Tongqiao Formula in Improving Unilateral Vestibular Labyrinth Destruction by Regulating Metabolism-neuroplasticity
Yu TIAN ; Hui LENG ; Rupeng QU ; Xianglong HAO ; Aiping WANG ; Lei SHI ; Zhongyuan QU ; Ye DONG ; Xiande MA ; Yangling HUANG
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(5):54-64
ObjectiveThis study aims to explore the mechanism by which Yishen Huoxue Tongqiao Formula improves metabolism-neuroplasticity and treats unilateral vestibular labyrinth destruction by regulating the metabolic balance of glutamate (Glu)/γ-aminobutyric acid (GABA). Methods48 Sprague-Dawley (SD) adult rats were randomly divided into the sham operation group, model group, Yishen Huoxue Tongqiao Formula groups with low, medium, and high doses (9.20, 18.39, 36.78 g·kg-1), and betahistine group (1.62 mg·kg-1). A unilateral vestibular labyrinth destruction (vestibular dysfunction) model was established by intratympanic injection of chloroform into the right ear, while the control group received intratympanic injection of normal saline. Drugs were administered once daily for seven consecutive days. During the period, behavioral tests were performed to evaluate the behaviors of rats after unilateral vestibular labyrinth destruction. Hematoxylin-eosin (HE) staining and Nissl staining were used to observe the neuronal morphology in the medial vestibular nucleus. Golgi staining was employed to assess the number of dendritic spines of neurons in the medial vestibular nucleus. Ultra-performance liquid chromatography-tandem mass spectrometry (LC-ESI-MS/MS) was utilized to detect Glu/GABA. Immunofluorescence and immunohistochemistry were used to detect the expressions of neuronal nuclei (NeuN), growth-associated protein 43 (GAP-43), and glial fibrillary acidic protein (GFAP). Western blot and real-time fluorescent quantitative polymerase chain reaction (Real-time PCR) were applied to determine the expressions of glutamate-immunoreactive (Glu-IR), GABA, GFAP, postsynaptic density protein 95 (PSD-95), and GAP-43. ResultsCompared with the sham operation group, the model group presented with head deviation, balance disorder, increased tail suspension score, nuclear consolidation of medial vestibular nerve neurons, and decreased Nissl bodies (P<0.01). The number of dendritic spines in neurons and NeuN-positive cells decreased. The content of Glu decreased. The content of GABA increased (Glu/GABA decreased). The expression of GAP-43 was down-regulated, and GFAP was up-regulated (P<0.05, P<0.01). The expressions of Glu-IR, PSD-95, and GAP-43 proteins, as well as Glu-IR mRNA decreased, while the expressions of GABA and GFAP proteins and mRNA increased (P<0.05, P<0.01). Compared with those in the model group, the head deviation, imbalanced behavior, and tail suspension scores in each treatment group decreased, with alleviated neuronal injury and recovered Nissl bodies (P<0.01). The number of dendritic spines of neurons increased, and the number of NeuN-positive cells rebounded. The content of Glu increased, and the content of GABA decreased (Glu/GABA increased). GFAP was down-regulated, and GAP-43 was up-regulated (P<0.05, P<0.01). The expressions of Glu-IR, PMD-95, and GAP-43 proteins, as well as Glu-IR mRNA increased, while the expressions of GABA and GFAP proteins and mRNA decreased. The effect was more significant in the high-dose group (P<0.01). ConclusionThe Yishen Huoxue Tongqiao Formula can alleviate vestibular dysfunction, and its mechanism may be associated with regulating the metabolic balance of Glu/GABA, mitigating neural damage, improving synaptic plasticity (promoting GAP-43 expression and inhibiting GFAP expression), and facilitating vestibular compensation.
2.Advances in Diabetic Peripheral Neuropathy Treatment by Traditional Chinese Medicine Based on Cellular Senescence: A Review
Qixian MA ; Shiyu HAN ; Hui HUANG ; Jing TIAN ; Xu HAN ; Qingguang CHEN ; Hao LU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(5):322-330
Diabetic Peripheral Neuropathy (DPN) is one of the most common and harmful complications of type 2 diabetes. DPN's pathogenesis include high blood sugar-induced oxidative stress, inflammation, and mitochondrial dysfunction. These factors are combined to damage nerve fibers, leading to sensory issues, pain, and numbness. Through a coordinated effect, these factors trigger nerve fiber damage and lead to sensory abnormalities, pain and numbness in limbs, and other symptoms, seriously restricting patients' activities of daily living and mobility. Recent research highlights that cellular senescence plays a critical role in DPN. Cellular senescence is manifested by the loss of cell proliferation ability, and further aggravates nerve damage via oxidative stress, mitochondrial dysfunction, autophagy impairment, inflammatory reaction, and other mechanisms, accelerating DPN occurrence and progression. In terms of medical treatment, current methods focus on blood sugar control, pain relief medicine, and microcirculation improvement, while no therapy has been developed based on cellular senescence. In contrast, traditional Chinese medicine (TCM) shows a unique advantage in DPN prevention and treatment via cellular senescence modulation. TCM emphasizes a holistic approach, as well as syndrome differentiation and treatment, effective in anti-aging and nerve damage repair. Recent studies show that TCM active ingredients, including puerarin, ginsenosides, and berberine, can reduce inflammation, oxidative stress, and apoptosis via signaling pathway regulation, thereby slowing cellular senescence to alleviate nerve damage. Furthermore, TCM compounds such as Buyang Huanwutang, Taohong Siwutang, and Huangqi Guizhi Wuwutang exert synergistic effects on cellular senescence-related pathways to improve nerve health and reduce DPN clinical symptoms. Therefore, this paper reviews the literature related to the interaction between cellular senescence and DPN from the perspective of cellular senescence, summarizing the mechanism of DPN and TCM intervention strategies.
3.Herbal Textual Research on Dioscoreae Hypoglaucae Rhizoma, Dioscoreae Spongiosae Rhizoma, Smilacis Chinae Rhizoma and Smilacis Glabrae Rhizoma in Famous Classical Formulas
Li LU ; Yichen YANG ; Erhuan WANG ; Hui CHANG ; Li AN ; Shibao WANG ; Cunde MA ; Zhilai ZHAN
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(5):218-247
This article systematically reviews and verifies the medicinal materials of Dioscoreae Hypoglaucae Rhizoma(DHR), Dioscoreae Spongiosae Rhizoma(DSR), Smilacis Chinae Rhizoma(SCR) and Smilacis Glabrae Rhizoma(SGR) from the aspects of name, origin, producing area, quality, harvesting, processing and efficacy by consulting historical literature, in order to provide reference for the development and utilization of famous classical formulas containing the four medicinal materials. DHR, DSR, SCR and SGR have a long history of application as medicinal materials. However, due to their similar growth environment and medicinal properties, as well as their functions of promoting dampness, dispelling wind and removing numbness, there have been instances of homonymous foreign objects and homonymous synonyms throughout history, resulting in confusion of the origin. Therefore, it is necessary to conduct comparative analysis and systematic research for clarifying the historical development and changes of the four, in order to provide a basis for safe and effective medication. According to research, Bixie was first recorded in Shennong Bencaojing and has been historically known as Baizhi, Chijie, Zhumu, and other aliases. From ancient times to the mid-20th century, there has always been a situation where the rhizomes of Dioscorea plants and Smilax plants, and even the rhizomes of Heterosmilax plants, were mixed together to be used as medicinal herbs for Bixie. However, since the Tang dynasty, it has been clearly advocated that the rhizomes of Dioscorea plants have excellent quality and have been the mainstream throughout history. The 2020 edition of Chinese Pharmacopoeia categorized it into two types of medicinal herbs(DHR and DSR). Among them, the origin of DHR is the dry rhizomes of Dioscorea hypoglauca, and the origins of DSR are the dry rhizomes of D. spongiosa and D. futschauensis. In ancient times, due to different types, the corresponding production areas of DHR and DSR were also different. Nowadays, They are mainly produced in the southern region of the Yangtze River. Since the Tang dynasty, the quality of Bixie has been characterized by its white color and soft nature. In modern times, it has been summarized that those with white color, large and thin pieces, powdery texture, tough and elastic texture, and neat and unbreakable are the best. The harvesting times of DHR and DSR are in spring or autumn, with the best quality harvested in autumn. The mainstream processing methods of them are slicing and then using the raw products or wine-processed products. SCR was first recorded in Mingyi Bielu and has been known as Jinganggen, Tielingjiao, Tieshuazi, and other aliases in history. The mainstream source is the dry rhizomes of Smilax china in the past dynasties, with the best quality being those that are tough and rich in powder. The harvesting time is from the late autumn to the following spring, and the main processing method throughout history has been slicing for raw use. SGR was first recorded under the item of Yuyuliang in Variorum of Shennong's Classic of Materia Medica. It was listed as an independent medicinal material from Bencao Gangmu. In history, there were such aliases as Cao Yuyuliang, Lengfantuan, Xianyiliang, Tubixie, etc. The main source of the past dynasties was dry rhizomes of S. glabra. In history, there have also been instances of multiple plants belonging to the same genus, and even cases of mixing the rhizomes of plants in the genus Heterosmilax. It is mainly produced in Guangdong, Hunan, Hubei, Zhejiang, Sichuan, Anhui and other regions, its quality has been summarized as large in size, powdery in texture, with few veins, and light brown in cross-section since modern times. The harvesting time is in spring or autumn, and the main processing method throughout history has been slicing for raw use. DHR, DSR, SCR and SGR all have the effects of promoting dampness, dispelling wind, relieving rheumatism and detoxifying. However, their detoxification abilities are ranked as follows:SGR>SCR>Bixie(DHR and DSR). Especially for the treatment of limb spasms, arthralgia and myalgia, scrofula, and scabies caused by syphilis and mercury poisoning, SGR has a unique effect. Based on the research results, DHR is recommended to develop the famous classical formulas containing Bixie as the first choice for medicinal herbs. It should be harvested in autumn, sliced thinly while fresh, and processed according to the requirements of the famous classical formulas, without any requirements for raw use. Selecting the rhizomes of S. china, harvested in late autumn, and thinly sliced while fresh. If there are no special processing requirements in the formulas, use it raw. Selecting the rhizomes of S. glabra, it is harvested in autumn and thinly sliced while fresh. If there are no special processing requirements in the formulas, raw products can be used.
4.Ameliorating effects of tetrahydrocurcumin and its nano-preparations on lipopolysaccharide-induced depression in mice
Hui Tan ; Yuanping Li ; Jingyuan Meng ; Tengteng Ma ; Yan Yang ; Zhengmao Yang ; Jiaqing Ma ; Jianping Xie ; Ying Guo
Acta Universitatis Medicinalis Anhui 2025;60(1):79-86
Objective :
To investigate the antidepressant effects and the underlying mechanisms of tetrahydrocurcumin(THC) and its nanoparticle formulation(THCN).
Methods :
Forty-six male ICR mice were randomly divided into Con group, LPS group, THC group, THCN group and SER group. A mouse depression model was established by intraperitoneal administration of LPS. The anxiety and depression-like behaviors of mice were evaluated by open field test(OFT) and forced swimming test(FST). Myelin staining was applied to assess the extent of demyelination in the prefrontal cortex of the mice. The prefrontal cortex and hippocampus were further examined for the expression levels of glial fibrillary acidic protein(GFAP) and Toll-like receptor 4(TLR4) through quantitative immunofluorescence assays.
Results :
Compared with the Con group, the LPS group showed increased anxiety-like and depressive-like behaviors in both the long-term and short-term experiments(P<0.05); the degree of demyelination increased in the LPS group of the long-term experiment(P<0.01); the expression of GFAP was reduced in the LPS group of the short-term experiment(P<0.01), while the expression of TLR4 increased(P<0.05); the expression of TLR4 decreased in the THC group(P<0.01); the expression of GFAP in the prefrontal cortex of the THCN group was reduced(P<0.01), while the expression of TLR4 increased(P<0.05). Compared with the LPS group, the THC group showed reduced depressive-like behaviors in the long-term experiment(P<0.05), while the anxiety-like and depressive-like behaviors of the THCN group and the SER group were reduced(P<0.05), and the anxiety-like and depressive-like behaviors of the THC group and the THCN group were reduced in the short-term experiment(P<0.05); the degree of demyelination was reduced in the THC group, THCN group and SER group in the long-term experiment(P<0.05); the expression of GFAP increased in the THC group of the short-term experiment(P<0.05), while the expression of TLR4 was reduced(P<0.05), and the expression of GFAP increased in the THCN group(P<0.05). Compared with the THC group, the THCN group and the SER group showed reduced anxiety-like behaviors in the long-term experiment(P<0.05); the expression of GFAP in the prefrontal cortex of the THCN group was reduced in the short-term experiment(P<0.05), while the expression of TLR4 in the hippocampal DG area increased in the short-term experiment(P<0.01).
Conclusion
Tetrahydrocurcumin and its nanoparticle formulation both exert significant ameliorative effects on depression-like behaviors and demyelination in mice induced by lipopolysaccharide. The antidepressant mechanism of THC appears to be mediated through the down-regulation of TLR4 and the up-regulation of GFAP. The mechanism underlying the antidepressant action of THCN seems predominantly focused on the enhancement of GFAP expression.
5.Precise application of O-arm navigation system in thoracolumbar fractures with developmental pedicle stenosis
Lintao SU ; Jianfeng JIANG ; Jun MA ; Liangliang HUANG ; Changyu LEI ; Yaozheng HAN ; Hui KANG
Chinese Journal of Tissue Engineering Research 2025;29(9):1855-1862
BACKGROUND:For thoracolumbar spine fractures with developmental stenosis of the vertebral arch,accurate nail placement is difficult using traditional fluoroscopy-assisted techniques.O-arm navigation assistance systems offer higher precision in general vertebral arch nail placement,but there is scarce literature on the application of O-arm navigation-assisted nail placement in thoracolumbar spine fractures with developmental stenosis of the vertebral arch both domestically and abroad. OBJECTIVE:To explore the accuracy of percutaneous vertebral arch nail placement assisted by O-arm navigation in patients with thoracolumbar spine fractures complicated by developmental stenosis of the vertebral arch. METHODS:A retrospective analysis was conducted on 53 patients who underwent percutaneous vertebral arch screw fixation surgery at Department of Orthopedics,General Hospital of Central Theater Command of PLA for thoracolumbar spine fractures complicated by developmental stenosis of the vertebral arch from January 2021 to March 2023.Totally 208 cases of vertebral arch developmental stenosis were found(cases with multiple vertebral arch developmental stenosis were counted separately).Based on the surgical approach,the patients were divided into two groups:O-arm navigation group(n=98)and C-arm fluoroscopy group(n=110).Postoperative imaging data were compared between the two groups,including anatomical perforation score,functional perforation score,actual vs.expected nail trajectory in the horizontal plane,and sagittal plane angle differences. RESULTS AND CONCLUSION:(1)There was no significant difference in the narrowest width of the pedicle isthmus(pow)between the two groups of patients(P>0.05).The proportions of different degrees of narrowing(mild:6 mm≤pow<7 mm,moderate:5 mm≤pow<6 mm,severe:pow<5 mm)were also not significantly different between the two groups(P>0.05).(2)The overall grade and scores of anatomical perforation and functional perforation were lower in the O-arm group compared to the C-arm group,and these differences were statistically significant(P<0.001).In terms of the angular deviation between the actual and planned screw trajectories,the O-arm group had smaller deviations,and these differences were statistically significant(P<0.05).(3)In the mild and moderate narrowing groups,the O-arm group showed significant advantages in anatomical perforation,functional perforation,and angular deviation between actual and planned screw trajectories,and these differences were statistically significant(P<0.001).(4)The O-arm group demonstrated better performance in anatomical perforation and functional perforation,especially in the T12-L2 segment,with more significant advantages.Additionally,the O-arm group had better angular deviations in actual and planned screw trajectories in all segments compared to the C-arm group.(5)Therefore,the use of O-arm navigation-assisted percutaneous screw placement for the treatment of thoracolumbar fractures with developmental pedicle isthmal narrowing provides higher accuracy and safer surgery.
6.The introduction on standards system of the pharmaceutical packaging materials in the Chinese Pharmacopoeia 2025 Edition
CHEN Lei ; YU Hui ; WANG Yan ; ZHANG Jun ; MA Shuangcheng
Drug Standards of China 2025;26(1):067-076
The standard of Pharmaceutical packaging materials is an important part of the Chinese Pharmacopoeia. This article focuses on working background, general idea, working process, main framework, and its role and significance of the pharmaceutical packaging materials standards system in the Chinese Pharmacopoeia 2025 Edition, which can contribute to accurately understand and utilize the standards in the Chinese Pharmacopoeia 2025 Edition.
7.Exposure to indoor chemical pollutants and associated health risk assessment of residents in Shijiazhuang city
Siwei ZHANG ; Yishan DING ; Xixi DU ; Fengge CHEN ; Junwang TONG ; Hui MA
Journal of Environmental and Occupational Medicine 2025;42(6):706-711
Background Indoor air quality directly affects people's health, especially the impact of chemical pollutants in residential indoor air on children and the elderly is more significant. Objective To understand the pollution status of common chemical pollutants in residential indoor air in Shijiazhuang, evaluate the health risks of chemical pollutants to school-age children and the elderly, and provide reference for controlling indoor pollution in residential environment. Methods Using stratified random sampling, a total of 60 households were selected from 2 urban areas and 1 surrounding rural area in Shijiazhuang City, specifically in July 2023 (non-heating season) and December 2023 (heating season), respectively. Relevant data was collected through on-site sampling [including CO, CO2, PM10, PM2.5, NO2, SO2, O3, ammonia, formaldehyde, benzene, toluene, xylene, total volatile organic compounds (TVOC), trichloroethylene, and tetrachloroethylene] and survey questionnaires. The pollutant concentrations were evaluated following the Standards for indoor air quality of GB/T 18883-2022, and the inhalation exposure risks of the target population were assessed based on the health risk assessment method. Results In the indoor air of the urban and rural residence in Shijiazhuang City, except for CO, NO2, SO2, toluene, and xylene, which did not exceed the standard limits, other pollutants showed varying degrees of exceedance. The non-qualified rates of PM10, PM2.5, and CO2 in the urban areas were higher than those in the rural areas (P < 0.05). The seasonal difference analysis showed that the non-qualified rates of PM2.5, PM10, CO2, trichloroethylene, and tetrachloroethylene in the urban areas were higher in the heating season than in the non-heating season (P<0.05); the non-qualified rates of ammonia and formaldehyde in the rural areas increased significantly in the non-heating season(P<0.05). The health risk assessment indicated that the maximum hazard quotient (HQ) of tetrachloroethylene for the elderly exceeded 1, while the HQ values for ammonia, formaldehyde, benzene, toluene, xylene, and trichloroethylene remained below 1 for either children or the elderly. For carcinogenic risks, the median carcinogenic risk (CR) of formaldehyde for school-aged children or the elderly fell within the range of 10−6-10−4, whereas the median CR values for benzene, trichloroethylene, and tetrachloroethylene were all below 10−6. Conclusion The primary indoor air pollutants exceeding the national standard limits in residential areas of Shijiazhuang City include CO2, PM10, TVOC, PM2.5, formaldehyde, ammonia, trichloroethylene, and tetrachloroethylene. The levels of these pollutants exhibit significant urban-rural and seasonal variations. Special attention should be paid to the non-carcinogenic risk of tetrachloroethylene to the elderly and the carcinogenic risks of formaldehyde to school-age children and the elderly.
8.Incremental effectiveness of two-dose of mumps-containing vaccine in chidren
Chinese Journal of School Health 2025;46(6):883-887
Objective:
To evaluate the incremental vaccine effectiveness (VE) of two dose of the mumps containing vaccine (MuCV) in chidren, so as to provide a basis for optimizing mumps immunization strategies.
Methods:
A 1∶2 frequency matched case-control study was conducted by using reported mumps cases in childcare centers or schools from Lu an, Hefei, Ma anshan and Huainan cities of Anhui Province from September 1, 2023 to June 30, 2024, as a case group(383 cases). And healthy children in the same classroom were selected as a control group(766 cases). The MuCV immunization histories of participants were collected to estimate the incremental VE of the second dose of MuCV against mumps. Group comparisons were performed using the Chi square test or t-test. For matched case-control pairs, the Cox regression model was employed to calculate the odds ratio (OR) with 95% confidence interval (CI) for two dose MuCV vaccination and to estimate the incremental vaccine effectiveness (VE).
Results:
There were no statistically significant differences between the case and control groups regarding gender, age, dosage of MuCV vaccination and the time interval since the last dose vaccination( χ 2/t=0.05, 0.20, 0.94, -0.02, P >0.05). The proportions of the case and control groups vaccinated with two doses of MuCV were 26.63% and 29.37%, respectively, and the overall incremental VE of the second dose of MuCV was 40.73% (95% CI=3.03%-63.77%, P <0.05). Subgroup analyses revealed that the incremental VE for children with a period of ≥1 year between the two doses of MuCV was 54.13% (95% CI=1.90%-78.56%, P <0.05), while for children with a period of <1 year, it was 30.63% (95% CI=-28.59%-62.58%, P >0.05). The incremental VE of the second dose of MuCV was 30.36% (95% CI=-25.95%-61.50%, P >0.05) in kindergarten children and 66.73% (95% CI=14.92%-86.99%, P <0.05) in elementary and secondary school students. The incremental VE was 28.78% (95% CI=-27.46%-60.21%, P >0.05) within five years of the last dose of MuCV vaccination and 66.07% (95% CI=-41.56%-91.87%, P >0.05) for vaccinations administered beyond five years.
Conclusions
The second dose of MuCV may offer additional protection for children; however, extending the interval between two dose of MuCV (<1 year) has shown limited incremental protective effects. Therefore, it is crucial to consider optimizing current immunization strategies for mumps.
9.Application of test of infant motor performance in the assessment of high-risk infants in Ma'anshan Area
Journal of Public Health and Preventive Medicine 2025;36(2):82-85
Objective To investigate common problems in motor development in high-risk infants and explore related influencing factors. Methods The high-risk infants who underwent the Test of Infant Motor Performance (TIMP) at the Early Childhood Development Department of Ma'anshan Maternal and Child Health Hospital from January 2021 to June 2023 were selected as the research subjects. Professional evaluation was conducted, and demographic information variables were collected. Finally, effective data of 1106 cases were obtained. The 2 test, t-test, or F-test were used to compare differences, and multiple linear regression analysis was used to explore the relevant factors influencing TIMP scores. Results According to the TIMP assessment, the incidence rates of developmental deviation and abnormality were 32.73% (362 /1 106) and 3.98% (44/1 106), respectively. Gender, birth weight, gestational age, test week age, and clinical impression were the influencing factors for TIMP scores (t-values=2.33, 2.26, -2.78, 23.35, and -8.48, all P<0.05). In the elicited items, except for the midline position of 15“ without vision and the upper limb defense of 30“, the scores of deviant and abnormal subjects were lower than those of excellent and good subjects, and the differences were statistically significant (P values<0.05). Conclusion The Chinese version of TIMP can reflect the level of exercise ability in high-risk infants and is suitable for early assessment of high-risk infants.
10.Impact of Toxoplasma gondii type I rhoptry protein 16 on programmed cell death ligand 1 expression and its binding to programmed cell death 1 in lung adenocarcinoma cells
Guangqi LI ; Yuning ZHOU ; Shaohan MA ; Mei TIAN ; Tiantian DANG ; Zhijun ZHAO
Chinese Journal of Schistosomiasis Control 2025;37(1):44-54
Objective To investigate the impact of Toxoplasma gondii type I, II and III rhoptry protein 16 (ROP16) on programmed cell death ligand 1 (PD-L1) expression in lung adenocarcinoma cells, and to examine the effects of T. gondii type I ROP16 protein on the relative PD-L1 expression, the relative PD-L1 distribution on the cell membrane surface, and the binding of programmed cell death 1 (PD-1) to PD-L1 in lung adenocarcinoma cells. Methods Lentiviral vectors overexpressing T. gondii type I, II and III ROP16 proteins were generated, and transfected into the human lung adenocarcinoma A549 cell line. A549 cells were used as a blank control group, and A549 cells transfected with an empty lentiviral expression vector were used as a negative control group, while A549 cells transfected with lentiviral vectors overexpressing T. gondii type I, II and III ROP16 proteins served as experimental groups. Stably transfected cells were selected with puromycin and verified using Western blotting, quantitative real-time PCR (RT-qPCR), and immunofluorescence assays. The PD-L1 expression was quantified at translational and transcriptional levels using Western blotting and RT-qPCR assays in A549 cells in the five groups, and the relative PD-L1 distribution was detected on the A549 cell membrane surface using flow cytometry. In addition, the effect of T. gondii type I ROP16 protein on the PD-1/PD-L1 binding was measured in A549 cells using enzyme-linked immunosorbent assay (ELISA). Results The relative ROP16 protein expression was 0, 0, 1.546 ± 0.091, 1.822 ± 0.047 and 2.334 ± 0.089 in the blank control group, negative control group, and the T. gondii type I, II and III ROP16 protein overexpression groups (F = 1 339.00,P < 0.001), and the relative ROP16 mRNA expression was 2.153 ± 0.949, 2.436 ± 1.614, 14.343 ± 0.020, 12.577 ± 0.285 and 15.090 ± 0.420 in the blank control group, negative control group and the T. gondii type I, II and III ROP16 protein overexpression groups, respectively (F = 483.50,P < 0.001). The ROP16 expression was higher in the T. gondii type I, II and III ROP16 protein overexpression groups than in the blank control group at both translational and transcriptional levels (allP values < 0.001). Immunofluorescence assay revealed that T. gondii type I, II and III ROP16 proteins were predominantly localized in A549 cell nuclei. Western blotting showed that the relative PD-L1 protein expression was 0.685 ± 0.109, 0.589 ± 0.114, 1.007 ± 0.117, 0.572 ± 0.151, and 0.426 ± 0.116 in the blank control group, negative control group, and the T. gondii type I, II and III ROP16 protein overexpression groups (F = 9.46,P < 0.05), and RT-qPCR assay quantified that the relative PD-L1 mRNA expression was 1.012 ± 0.190, 1.281 ± 0.465, 1.950 ± 0.175, 0.889 ± 0.251, and 0.230 ± 0.192 in the blank control group, negative control group, and the T. gondii type I, II and III ROP16 protein overexpression groups (F = 14.18,P < 0.05). The PD-L1 expression was higher in the T. gondii type IROP16 protein overexpression group than in the blank control group at both translational and transcriptional levels (both P values < 0.05). Flow cytometry detected that the relative distributions of PD-L1 protein were (10.83 ± 0.60)%, (11.23 ± 0.20)%, and (14.61 ± 0.50)% on the A549 cell membrane surface (F = 28.31, P < 0.05), and the relative distribution of PD-L1 protein was higher in the T. gondii type IROP16 protein overexpression group than in the blank control group and negative control group (both P values < 0.001). ELISA measured significant differences in the absorbance (A) value among the T. gondii type IROP16 protein overexpression group, the blank control group and the negative control group if the concentrations of the recombinant PD-1 protein were 0.04 (F = 10.45, P < 0.05), 0.08 μg/mL (F = 11.68, P < 0.05) and 0.12 μg/mL (F = 52.68, P < 0.05), and the A value was higher in the T. gondii type IROP16 protein overexpression group than in the blank control group and the negative control group (both P values < 0.05), indicating that T. gondii type IROP16 protein promoted the PD-L1/PD-1 binding in A549 cells in a concentration-dose manner. Conclusions T. gondii type IROP16 protein overexpression may up-regulate PD-L1 expression in A549 cells at both transcriptional and translational levels and the relative PD-L1 distribution on the A549 cell membrane surface, and affect the PD-1/PD-L1 binding in a concentration-dependent manner.


Result Analysis
Print
Save
E-mail