[摘 要] 目的：观察白细胞介素-37（IL-37）在乳腺癌患者的表达变化对CD8+ T细胞活性的影响。方法：纳入2020年7月至2022年9月在新乡医学院第一附属医院就诊的46例乳腺癌患者、24例乳腺良性肿瘤患者、20例对照者。采集外周血，分离血浆和外周血单个核细胞（PBMC），收集接受手术治疗的乳腺癌患者肿瘤组织和癌旁组织，分离组织中肿瘤浸润淋巴细胞（TIL），纯化CD8+ T细胞。ELISA法检测IL-37、可溶型单免疫球蛋白IL-1受体相关蛋白（SIGIRR）表达，实时定量PCR法检测组织中IL-37 mRNA，流式细胞术检测CD8+ T细胞中IL-18受体α链（IL-18Rα）和SIGIRR表达。外源性IL-37刺激纯化的CD8+ T细胞，与乳腺癌细胞系MCF-7共培养，通过测定乳酸脱氢酶水平计算靶细胞死亡比例，ELISA法检测上清中穿孔素、颗粒酶B、干扰素-γ（IFN-γ）、肿瘤坏死因子-α（TNF-α）水平。结果：乳腺癌患者血浆IL-37水平高于乳腺良性肿瘤患者[（554.17 ± 96.63）pg/mL vs （499.52 ± 78.66）pg/mL，P = 0.020]和对照者[（483.97 ± 47.23）pg/mL，P = 0.003]。乳腺癌患者肿瘤组织中IL-37 mRNA相对表达量高于癌旁组织[（1.88 ± 0.21） vs （1.00 ± 0.53）pg/mL，P < 0.001]。外周血IL-18Rα+ CD8+细胞比例、SIGIRR+ CD8+细胞比例、血浆可溶型SIGIRR水平在乳腺癌患者、乳腺良性肿瘤患者、对照者之间的差异无统计学意义（均P > 0.05）。CD8+ TIL表达IL-18Rα和SIGIRR的比例在肿瘤组织和癌旁组织之间的差异无统计学意义（P > 0.05）。重组人IL-37刺激后，CD8+ T细胞诱导靶细胞死亡比例、上清中IFN-γ和TNF-α水平在直接接触和间接接触共培养系统中均低于无刺激（均P < 0.05）。在直接接触共培养系统中，IL-37刺激后上清中穿孔素和颗粒酶B水平均低于无刺激（均P < 0.001），但在间接接触共培养系统中，上清中穿孔素和颗粒酶B水平在无刺激和IL-37刺激之间的差异无统计学意义（均P > 0.05）。结论：乳腺癌患者中IL-37水平升高可能参与诱导外周血和肿瘤微环境中CD8+ T细胞功能衰竭。

Objective:To explore the etiology of primary amenorrhea,analyze its clinical features,and improve the level of clinical diagnosis and treatment.Methods:A retrospective analysis was conducted on the clinical data of 75 patients with primary amenorrhea who attended the gynecological endocrine clinic of Nanjing Women and Children's Healthcare Hospital from January 2020 to September 2022,and the etiology of their diagnosis was ana-lyzed.Results:aChromosomal karyotype analysis showed 14 cases(18.67％)of abnormal karyotypes,including 2 cases of autosomal abnormalities,7 cases of sex chromosome abnormalities,and 5 cases of sexual differentia-tion abnormalities.②According to reproductive hormones,there were 15 cases(20.00％)of hypogonadotropic a-menorrhea and 18 cases(24.00％)of hypergonadotropic amenorrhea.③By the location of the cause,utero-inferi-or genital tract amenorrhea was the most common in 23 cases(30.67％),including 20 cases of mayer-rokitan-sky-küster-hauser syndrome(MRKH syndrome)and 3 cases of androgen insensitivity syndrome(AIS).It was followed by ovarian amenorrhea in 18 cases(24.00％)including 6 cases of Turners syndrome,10 cases of 46,XX simple gonadal hypoplasia and 2 cases of 46,XY gonadal hypoplasia(Swyer syndrome);hypothalamic-pituitary amenorrhea in 17 cases(22.67％),including idiopathic hypogonadotropic hypogonadism(IHH)in 9 cases,Kall-mann s syndrome in 4 cases,and hyperprolactinemia in 4 cases;and also amenorrhea with hyperandrogenic mani-festations in 17 cases(22.67％),with a higher number of suspicious polycystic ovary syndrome in 16 cases.Con-clusions:Primary amenorrhea has a variety of etiologies and clinical features,and requires examination of sex hormones and pelvic ultrasound from the lower genital tract up to the central nervous system to find the site of the etiology and to optimize the diagnostic process;in the case of hypogonadotropic amenorrhea,chromosomal test is necessary.

Objective:To explore the etiology of primary amenorrhea,analyze its clinical features,and improve the level of clinical diagnosis and treatment.Methods:A retrospective analysis was conducted on the clinical data of 75 patients with primary amenorrhea who attended the gynecological endocrine clinic of Nanjing Women and Children's Healthcare Hospital from January 2020 to September 2022,and the etiology of their diagnosis was ana-lyzed.Results:aChromosomal karyotype analysis showed 14 cases(18.67％)of abnormal karyotypes,including 2 cases of autosomal abnormalities,7 cases of sex chromosome abnormalities,and 5 cases of sexual differentia-tion abnormalities.②According to reproductive hormones,there were 15 cases(20.00％)of hypogonadotropic a-menorrhea and 18 cases(24.00％)of hypergonadotropic amenorrhea.③By the location of the cause,utero-inferi-or genital tract amenorrhea was the most common in 23 cases(30.67％),including 20 cases of mayer-rokitan-sky-küster-hauser syndrome(MRKH syndrome)and 3 cases of androgen insensitivity syndrome(AIS).It was followed by ovarian amenorrhea in 18 cases(24.00％)including 6 cases of Turners syndrome,10 cases of 46,XX simple gonadal hypoplasia and 2 cases of 46,XY gonadal hypoplasia(Swyer syndrome);hypothalamic-pituitary amenorrhea in 17 cases(22.67％),including idiopathic hypogonadotropic hypogonadism(IHH)in 9 cases,Kall-mann s syndrome in 4 cases,and hyperprolactinemia in 4 cases;and also amenorrhea with hyperandrogenic mani-festations in 17 cases(22.67％),with a higher number of suspicious polycystic ovary syndrome in 16 cases.Con-clusions:Primary amenorrhea has a variety of etiologies and clinical features,and requires examination of sex hormones and pelvic ultrasound from the lower genital tract up to the central nervous system to find the site of the etiology and to optimize the diagnostic process;in the case of hypogonadotropic amenorrhea,chromosomal test is necessary.

Objective:To explore the etiology of primary amenorrhea,analyze its clinical features,and improve the level of clinical diagnosis and treatment.Methods:A retrospective analysis was conducted on the clinical data of 75 patients with primary amenorrhea who attended the gynecological endocrine clinic of Nanjing Women and Children's Healthcare Hospital from January 2020 to September 2022,and the etiology of their diagnosis was ana-lyzed.Results:aChromosomal karyotype analysis showed 14 cases(18.67％)of abnormal karyotypes,including 2 cases of autosomal abnormalities,7 cases of sex chromosome abnormalities,and 5 cases of sexual differentia-tion abnormalities.②According to reproductive hormones,there were 15 cases(20.00％)of hypogonadotropic a-menorrhea and 18 cases(24.00％)of hypergonadotropic amenorrhea.③By the location of the cause,utero-inferi-or genital tract amenorrhea was the most common in 23 cases(30.67％),including 20 cases of mayer-rokitan-sky-küster-hauser syndrome(MRKH syndrome)and 3 cases of androgen insensitivity syndrome(AIS).It was followed by ovarian amenorrhea in 18 cases(24.00％)including 6 cases of Turners syndrome,10 cases of 46,XX simple gonadal hypoplasia and 2 cases of 46,XY gonadal hypoplasia(Swyer syndrome);hypothalamic-pituitary amenorrhea in 17 cases(22.67％),including idiopathic hypogonadotropic hypogonadism(IHH)in 9 cases,Kall-mann s syndrome in 4 cases,and hyperprolactinemia in 4 cases;and also amenorrhea with hyperandrogenic mani-festations in 17 cases(22.67％),with a higher number of suspicious polycystic ovary syndrome in 16 cases.Con-clusions:Primary amenorrhea has a variety of etiologies and clinical features,and requires examination of sex hormones and pelvic ultrasound from the lower genital tract up to the central nervous system to find the site of the etiology and to optimize the diagnostic process;in the case of hypogonadotropic amenorrhea,chromosomal test is necessary.

Objective:To explore the etiology of primary amenorrhea,analyze its clinical features,and improve the level of clinical diagnosis and treatment.Methods:A retrospective analysis was conducted on the clinical data of 75 patients with primary amenorrhea who attended the gynecological endocrine clinic of Nanjing Women and Children's Healthcare Hospital from January 2020 to September 2022,and the etiology of their diagnosis was ana-lyzed.Results:aChromosomal karyotype analysis showed 14 cases(18.67％)of abnormal karyotypes,including 2 cases of autosomal abnormalities,7 cases of sex chromosome abnormalities,and 5 cases of sexual differentia-tion abnormalities.②According to reproductive hormones,there were 15 cases(20.00％)of hypogonadotropic a-menorrhea and 18 cases(24.00％)of hypergonadotropic amenorrhea.③By the location of the cause,utero-inferi-or genital tract amenorrhea was the most common in 23 cases(30.67％),including 20 cases of mayer-rokitan-sky-küster-hauser syndrome(MRKH syndrome)and 3 cases of androgen insensitivity syndrome(AIS).It was followed by ovarian amenorrhea in 18 cases(24.00％)including 6 cases of Turners syndrome,10 cases of 46,XX simple gonadal hypoplasia and 2 cases of 46,XY gonadal hypoplasia(Swyer syndrome);hypothalamic-pituitary amenorrhea in 17 cases(22.67％),including idiopathic hypogonadotropic hypogonadism(IHH)in 9 cases,Kall-mann s syndrome in 4 cases,and hyperprolactinemia in 4 cases;and also amenorrhea with hyperandrogenic mani-festations in 17 cases(22.67％),with a higher number of suspicious polycystic ovary syndrome in 16 cases.Con-clusions:Primary amenorrhea has a variety of etiologies and clinical features,and requires examination of sex hormones and pelvic ultrasound from the lower genital tract up to the central nervous system to find the site of the etiology and to optimize the diagnostic process;in the case of hypogonadotropic amenorrhea,chromosomal test is necessary.

Objective:To explore the etiology of primary amenorrhea,analyze its clinical features,and improve the level of clinical diagnosis and treatment.Methods:A retrospective analysis was conducted on the clinical data of 75 patients with primary amenorrhea who attended the gynecological endocrine clinic of Nanjing Women and Children's Healthcare Hospital from January 2020 to September 2022,and the etiology of their diagnosis was ana-lyzed.Results:aChromosomal karyotype analysis showed 14 cases(18.67％)of abnormal karyotypes,including 2 cases of autosomal abnormalities,7 cases of sex chromosome abnormalities,and 5 cases of sexual differentia-tion abnormalities.②According to reproductive hormones,there were 15 cases(20.00％)of hypogonadotropic a-menorrhea and 18 cases(24.00％)of hypergonadotropic amenorrhea.③By the location of the cause,utero-inferi-or genital tract amenorrhea was the most common in 23 cases(30.67％),including 20 cases of mayer-rokitan-sky-küster-hauser syndrome(MRKH syndrome)and 3 cases of androgen insensitivity syndrome(AIS).It was followed by ovarian amenorrhea in 18 cases(24.00％)including 6 cases of Turners syndrome,10 cases of 46,XX simple gonadal hypoplasia and 2 cases of 46,XY gonadal hypoplasia(Swyer syndrome);hypothalamic-pituitary amenorrhea in 17 cases(22.67％),including idiopathic hypogonadotropic hypogonadism(IHH)in 9 cases,Kall-mann s syndrome in 4 cases,and hyperprolactinemia in 4 cases;and also amenorrhea with hyperandrogenic mani-festations in 17 cases(22.67％),with a higher number of suspicious polycystic ovary syndrome in 16 cases.Con-clusions:Primary amenorrhea has a variety of etiologies and clinical features,and requires examination of sex hormones and pelvic ultrasound from the lower genital tract up to the central nervous system to find the site of the etiology and to optimize the diagnostic process;in the case of hypogonadotropic amenorrhea,chromosomal test is necessary.

Objective:To explore the etiology of primary amenorrhea,analyze its clinical features,and improve the level of clinical diagnosis and treatment.Methods:A retrospective analysis was conducted on the clinical data of 75 patients with primary amenorrhea who attended the gynecological endocrine clinic of Nanjing Women and Children's Healthcare Hospital from January 2020 to September 2022,and the etiology of their diagnosis was ana-lyzed.Results:aChromosomal karyotype analysis showed 14 cases(18.67％)of abnormal karyotypes,including 2 cases of autosomal abnormalities,7 cases of sex chromosome abnormalities,and 5 cases of sexual differentia-tion abnormalities.②According to reproductive hormones,there were 15 cases(20.00％)of hypogonadotropic a-menorrhea and 18 cases(24.00％)of hypergonadotropic amenorrhea.③By the location of the cause,utero-inferi-or genital tract amenorrhea was the most common in 23 cases(30.67％),including 20 cases of mayer-rokitan-sky-küster-hauser syndrome(MRKH syndrome)and 3 cases of androgen insensitivity syndrome(AIS).It was followed by ovarian amenorrhea in 18 cases(24.00％)including 6 cases of Turners syndrome,10 cases of 46,XX simple gonadal hypoplasia and 2 cases of 46,XY gonadal hypoplasia(Swyer syndrome);hypothalamic-pituitary amenorrhea in 17 cases(22.67％),including idiopathic hypogonadotropic hypogonadism(IHH)in 9 cases,Kall-mann s syndrome in 4 cases,and hyperprolactinemia in 4 cases;and also amenorrhea with hyperandrogenic mani-festations in 17 cases(22.67％),with a higher number of suspicious polycystic ovary syndrome in 16 cases.Con-clusions:Primary amenorrhea has a variety of etiologies and clinical features,and requires examination of sex hormones and pelvic ultrasound from the lower genital tract up to the central nervous system to find the site of the etiology and to optimize the diagnostic process;in the case of hypogonadotropic amenorrhea,chromosomal test is necessary.

Objective:To explore the etiology of primary amenorrhea,analyze its clinical features,and improve the level of clinical diagnosis and treatment.Methods:A retrospective analysis was conducted on the clinical data of 75 patients with primary amenorrhea who attended the gynecological endocrine clinic of Nanjing Women and Children's Healthcare Hospital from January 2020 to September 2022,and the etiology of their diagnosis was ana-lyzed.Results:aChromosomal karyotype analysis showed 14 cases(18.67％)of abnormal karyotypes,including 2 cases of autosomal abnormalities,7 cases of sex chromosome abnormalities,and 5 cases of sexual differentia-tion abnormalities.②According to reproductive hormones,there were 15 cases(20.00％)of hypogonadotropic a-menorrhea and 18 cases(24.00％)of hypergonadotropic amenorrhea.③By the location of the cause,utero-inferi-or genital tract amenorrhea was the most common in 23 cases(30.67％),including 20 cases of mayer-rokitan-sky-küster-hauser syndrome(MRKH syndrome)and 3 cases of androgen insensitivity syndrome(AIS).It was followed by ovarian amenorrhea in 18 cases(24.00％)including 6 cases of Turners syndrome,10 cases of 46,XX simple gonadal hypoplasia and 2 cases of 46,XY gonadal hypoplasia(Swyer syndrome);hypothalamic-pituitary amenorrhea in 17 cases(22.67％),including idiopathic hypogonadotropic hypogonadism(IHH)in 9 cases,Kall-mann s syndrome in 4 cases,and hyperprolactinemia in 4 cases;and also amenorrhea with hyperandrogenic mani-festations in 17 cases(22.67％),with a higher number of suspicious polycystic ovary syndrome in 16 cases.Con-clusions:Primary amenorrhea has a variety of etiologies and clinical features,and requires examination of sex hormones and pelvic ultrasound from the lower genital tract up to the central nervous system to find the site of the etiology and to optimize the diagnostic process;in the case of hypogonadotropic amenorrhea,chromosomal test is necessary.

Objective:To explore the etiology of primary amenorrhea,analyze its clinical features,and improve the level of clinical diagnosis and treatment.Methods:A retrospective analysis was conducted on the clinical data of 75 patients with primary amenorrhea who attended the gynecological endocrine clinic of Nanjing Women and Children's Healthcare Hospital from January 2020 to September 2022,and the etiology of their diagnosis was ana-lyzed.Results:aChromosomal karyotype analysis showed 14 cases(18.67％)of abnormal karyotypes,including 2 cases of autosomal abnormalities,7 cases of sex chromosome abnormalities,and 5 cases of sexual differentia-tion abnormalities.②According to reproductive hormones,there were 15 cases(20.00％)of hypogonadotropic a-menorrhea and 18 cases(24.00％)of hypergonadotropic amenorrhea.③By the location of the cause,utero-inferi-or genital tract amenorrhea was the most common in 23 cases(30.67％),including 20 cases of mayer-rokitan-sky-küster-hauser syndrome(MRKH syndrome)and 3 cases of androgen insensitivity syndrome(AIS).It was followed by ovarian amenorrhea in 18 cases(24.00％)including 6 cases of Turners syndrome,10 cases of 46,XX simple gonadal hypoplasia and 2 cases of 46,XY gonadal hypoplasia(Swyer syndrome);hypothalamic-pituitary amenorrhea in 17 cases(22.67％),including idiopathic hypogonadotropic hypogonadism(IHH)in 9 cases,Kall-mann s syndrome in 4 cases,and hyperprolactinemia in 4 cases;and also amenorrhea with hyperandrogenic mani-festations in 17 cases(22.67％),with a higher number of suspicious polycystic ovary syndrome in 16 cases.Con-clusions:Primary amenorrhea has a variety of etiologies and clinical features,and requires examination of sex hormones and pelvic ultrasound from the lower genital tract up to the central nervous system to find the site of the etiology and to optimize the diagnostic process;in the case of hypogonadotropic amenorrhea,chromosomal test is necessary.

Objective The objective of this study was to investigate the expression of interleukin-38(IL-38)in patients with breast cancer and its regulatory function to CD8+T cell activity.Methods 44 patients with breast cancer,25 patients with benign breast tumor,and 20 controls,who were treated in the First Affiliated Hospital of Xinxiang Medical University from July 2020 and Sep-tember 2022.Mononuclear cells from plasma and peripheral blood of all subjects were isolated,tumor-infiltrating lymphocytes from tumor tissues of breast cancer patients were isolated,and CD8+T cells were purified.IL-38 protein level in the plasma was measured by enzyme-linked immunosorbent assay(ELISA).The relative level of IL-38 mRNA in the tissue was semi-quantified by real-time quantitative PCR.Recombinant human IL-38 was used to stimulate CD8+T cells from peripheral blood and tumor tissue from patients with breast cancer.A co-culture system was established between CD8+T cells and breast cancer MCF-7 cell line.The percentage of target cell death was calculated by measuring lactate dehydrogenase level in the supernatants.The levels of perforin,granzyme B,inter-feron-γ and tumor necrosis factor-α(TNF-α)in the supernatants were measured by ELISA.The immune checkpoint molecules ex-pression in CD8+T cells were detected by flow cytometry.Results The levels of plasma IL-38 were significantly higher in patients with breast cancer(74.23±19.88 pg/mL)compared with in patients with benign breast tumor(62.87±16.27 pg/mL,P=0.018)and controls(61.77±12.75 pg/mL,P=0.013).The relative expression of IL-38 mRNA in tumor tissues was significantly higher than in para-tumor tissues(1.57±0.22 vs.1.00±0.18,P<0.001).The proportion of target cell death induced by peripheral and tumor-in-filtrating CD8+T cells,and the levels of perforin and granzyme B secretion in direct contact co-culture group were higher than those in indirect contact co-culture group(P<0.05).There were no significant differences of either interferon-γ or TNF-α levels between di-rect contact and indirect contact co-culture group(P>0.05).In the direct contact co-culture group,the levels of target cell death pro-portion,perforin,granzyme B,interferon-γ and TNF-α l in the IL-38 stimulation group were lower than those in the non-stimulation group(P<0.05).In the indirect contact co-culture group,the target of cell death proportion,interferon-γ and TNF-α the IL-38 stimulation group were also lower than those in the non-stimulation group(P<0.05).However,there were no statistical differences of either perforin or granzyme B levels between the IL-38 stimulation group and non-stimulation group within the indirect contact co-culture group(P>0.05).There were also no differences in the levels of immune checkpoint molecules in CD8+T cells between the non-stimulation group and the IL-38 stimulation group(P>0.05).Conclusion Highly expressed IL-38 in patients with breast cancer may be involved in inducing CD8+T cell functional failure.