IL-33/ST2 axis in systemic lupus erythematosus in relation to chronic kidney injury and disease activity

AIM: To elucidate the association between chronic kidney injury and interleukin-33 (IL-33; an alarmin)/suppression of tumorigencity 2 (ST2) in patients with systemic lupus erythematosus (SLE).METHODS: Serum levels of IL-33 and soluble ST2 (sST2) were assessed by ELISA in 50 SLE patients and 30 healthy controls (HC).RESULTS: The levels of IL-33 and sST2, and IL-33/sST2 ratio were significantly higher in SLE patients than those in the HC. The IL-33 and sST2 levels were positively associated with SLE disease activity index (SLEDAI), erythrocyte sedimentation rate (ESR), proteinuria and triglyceride, but negatively associated with complement C3. IL-33/sST2 ratio was positively associated with SLEDAI and estimated glomerular filtration rate (eGFR). Independent explanatory variables associated with high IL-33/sST2 included chronic kidney disease (CKD) staging and albumin (R²=0.442), especially CKD staging.CONCLUSION: Elevated serum sST2 and IL-33 levels in SLE patients are correlated with disease activity and risk factors of kidney injury. IL-33/sST2 ratio may serve as a potential biomarker for chronic kidney injury in SLE patients.     

Experimental Researches on Carcinogenesis or Tumorigenicity of MDCK Canine Kidney Cell(CKC) Lines and Analysis of Their Chromosome Karyotypes

The chromosomal number variations & structural aberrations of the MDCK cell line, primary feline or canine kidney cell(FKC or CKC) and Hela cell line were investigated and their karyotypes of conventional chromosome bands were analyzed. The carcinogenesis or tumorigenicity testing of these cell lines in about 232 nude mice and for colony formation in soft agarose and for haemagglutination under different concentration of plant lectins of these cells were carried out. Under the prerequisite that the incidence of cancer or tumor in negative-control nude mice inoculated subcutaneously with primary feline or canine kidney cell cultures purified in vitro at passage 3 was 0 (0/22) and 0 (0/10), respectively. The incidence of the progressively-growing malignant tumor(MT) in positive-control nude mice inoculated subcutaneously with Hela cell cultures of KB, X, or NM20/X strain was 10/10, 25/25 and 5/51, respectively. The results showed that the incidence of tumor in nude mice with tetrapioid YA strain of MDCK cell during 20 - 45 passages, with hypodiploid JB strain of MDCK cell on passage 25, with di-and hypoploid JC strain of MDCK cell during 2 - 15passages or with hypoploid M strain of MDCK cell during 9 - 27 passages was 28/58, 1/5, 4/18 and 0/31,respectively. The chromosomal analysis results showed that the ratio of difference in the rate of modal chromosome number between high (mcs + n) and lowest (mcs)passages was not more than 5 % - 15 % and the structure aberrations was generally 0 - 3%. These results proved that the genetic characteristics of chromosomal number of cell lines determines their tumorigenicity, but it is species-specific. MDCK line has tumorigenicity no matter what its chromosome karyotype is, at least it has very low tumorigenicity even when its modal chromosome number is hypoploid. The repeatedly frozen, thawed and split controls of tumorigenicity-positive cell lines(X strain of Hela, M strain of BHK-21, JA strain of Vero, YA strain of MDCK) have much lower tumorigenicity or are even non-carcinogenesis, and the repeatedly frozen, thawed and split controls of very low tumorigenicity cell lines (M or JC strain of MDCK) are certainly non-carcinogenic and never have increased tumorigenicity.It is thus evident that MDCK cell of M, JB or JC strain can be approved as substrate for the preparation of attenuated viral vaccines, but MDCK cell of YA strain can not be approved as substrate for the preparation of comattenuated viral vaccines. In summary, all strains of MDCK cell line have tunorigenicity, at least have low tumorigencity, never have non-cancinogenic MDCK, but very low tumorigenicity MDCK cell strains can certainly be used for the approval production of canine viral vaccines if the DNA content in viral cell cultures was remarkably decreased through conventional means in manufacturing process. Therefore, the master cell stock and working cell bank of MDCK line used for vaccine manufacture were established in China, which are free of infectious agents, and described with respect to cytogenetic characteristics and tumorigenicity. Tests showed that there were correlations among cell line chromosome number variations, anchorage independence in soft agarose, haemagglutination under plant lectins, and tumor-forming ability in nude mice, thus all the in vitro tests are economic, simple and reliable means for monitoring the tumor-forming ability of MDCK line in nude mice.     

国内外生产用传代细胞系致瘤性检验方法的比较与分析

对WHO 2013年公布的《动物细胞基质生产生物医药产品和细胞库特性的评价建议》及我国2015版《中华人民共和国兽药典》三部中涉及的细胞致瘤性检验相关要求进行了介绍、梳理和比较，并对我国兽用生物制品生产用细胞的致瘤性检验要求提供了进一步完善的建议。     

六种传代细胞系的致瘤性研究

为分析6种传代细胞系对裸鼠致瘤性的影响,将80只裸鼠随机分为8组,每组10只。处理组分为MDBK细胞组、OA3.Ts细胞组、Marc-145细胞组、McCoy细胞组、PK-15细胞组、ST细胞组,每种细胞培养物颈背部皮下注射10只裸鼠。阳性对照组BHK-21细胞培养物和阴性对照组鸡胚成纤维细胞培养物各注射10只裸鼠。在饲养5周和12周后对裸鼠剖检并进行病理组织学检测。结果发现,阳性对照BHK-21细胞组、McCoy细胞组、PK-15细胞组和Marc-145细胞组接种裸鼠后5周注射部位均形成肿块。对各处理组采集的肿块进行病理组织学检测,结果均为鳞状细胞癌细胞。对阳性对照BHK-21细胞组的一只裸鼠肺组织进行病理组织学检测发现有癌细胞的转移;阴性对照鸡胚成纤维细胞组、MDBK细胞组、OA3.Ts细胞组和ST细胞组小鼠均无肿块形成,接种后5周和12周分别进行剖检和病理组织学检测均未发现肿瘤细胞。由此可知,MDBK细胞、OA3.Ts细胞、ST细胞对裸鼠均不具有致瘤性;McCoy细胞、PK-15细胞、Marc-145细胞接种后可诱导裸鼠发生肿瘤,具有致瘤性;若使用致瘤性阳性细胞进行灭活疫苗生产时,应对灭活苗生产的工艺过程进行进一步的安全性评价。     

BHK-21细胞不同处理方式对裸鼠致瘤性的影响

为了分析BHK-21细胞对裸鼠致瘤性受何种因素的影响,我们分别用甲醛、二乙烯亚胺(BEI)、β-丙内酯(BPL)进行灭活和在-20℃进行冻融处理的BHK-21细胞接种裸鼠,通过病理组织形态学观察判定其是否存在致肿瘤的特性。结果显示,甲醛、BEI、BPL、冻融2次或3次分别处理的BHK-21细胞接种裸鼠,对裸鼠致瘤率为零。冻融1次的BHK-21细胞和107、105和103个BHK-21细胞分别接种裸鼠,致瘤率为100%。10个BHK-21细胞接种裸鼠,未见成瘤。本次试验表明,裸鼠检测BHK-21细胞致瘤性敏感性为103个细胞；冻融2次、甲醛、BEI、BPL处理,BHK-21细胞均丧失致瘤性；冻融1次不能使BHK-21细胞丧失致瘤性。