| Systemic lupus erythematosus(SLE), an autoimmunity disease with the prominent manifestations of immune inflammation, is clinically characterized by the generation of a variety of autoantibodies, represented by antinuclear antibodies, and multiple organ involvement, including the most commonly and seriously affected kidney. Kidney damage affected by SLE is called lupus nephritis(LN). According to the clinical manifestations, renal involvement exists in more than 70 % of SLE patients, and LN is the initial symptom in 20 % of SLE patients. Almost all SLE patients suffer from kidney damage according to the kidney pathological biopsy. The severity of the kidney lesions is an important factor that directly affects the prognosis of SLE. At present, pharmaceutical drug is still the main treatment strategy for LN. Glucocorticoid(GC) and immune-suppressors are mostly used drugs in the clinical treatment of LN. However, long-term use of GC and immune-suppressors may lead to additional infection and induce toxic or side effects including Cushing’s syndrome, osteoporosis, gastrointestinal tract stimulation, gonad inhibiting. Given serious threat of LN to public health, it is necessary to develop the new effective treatment strategy for LN.Chemokine is a kind of small molecule that can affect cell directional movement. As a basic protein, it is composed of 70~125 amino acids with the relative molecular mass being between 6 and 14 KD. More than 50 kinds of chemokines have been found so far and all of these chemokines can be classified as four different subtribes, namely C, CC, CXC and CX3 C. Chemokines and their receptors have been reported to function in regulating cell growth and other physiological process including immune system development and response, inflammation, tumor development and wound healing. Also, a growing number of studies show that chemokines involved in the pathogenesis of LN.Systemic lupus erythematosus(SLE), an autoimmunity disease with the prominent manifestations of immune inflammation, is clinically characterized by the generation of a variety of autoantibodies, represented by antinuclear antibodies, and multiple organ involvement, including the most commonly and seriously affected kidney. Kidney damage affected by SLE is called lupus nephritis(LN). The extent of renal damage can serve as an important factor that related to the prognosis of SLE patients. At present, pharmaceutical drug is still the main treatment strategy for LN. Glucocorticoid(GC) and immune-suppressors are mostly used drugs in the clinical treatment of LN. However, long-term use of GC and immune-suppressors may lead to additional infection and induce toxic or side effects including Cushing’s syndrome, osteoporosis, gastrointestinal tract stimulation, gonad inhibiting. Given serious threat of LN to public health, it is necessary to develop the new effective treatment strategy for LN.CXCL13, also known as B cell-attracting chemokine-1(BCA-1) is the main regulatory factor for B cells’ directional chemotaxis. A large number of studies have detected the high expression of CXCL13 in the serum of SLE patients and SLE animal models, which is related to the disease activity and severity. Further studies also indicate high expression of CXCL13 in the glomerular tissue of SLE mice. Therefore, CXCL13 is regarded as a biomarker and a potential therapeutic target of SLE. Thus, a hypothesis that using anti-CXCL13 monoclonal antibody to inhibit the activity of CXCL13 may be an effective method for treatment of SLE and LN is formulated. In the present study, treatment of CXCL13 monoclonal antibody to MRL/lpr mice was performed to explore the possibility of CXCL13 as a therapeutic target to LN.This study consists of three sections as follows:1. Alleviation of kidney injury in MRL/lpr mice by CXCL13 treatmentProteinuria is a common sign and objective indicator of kidney disease. Concentration of creatinine in serum depends on the filtration ability of glomerular, and high concentration level of creatinine in serum always means renal lesions.The present study divided 24 female MRL/lpr mice of 14 weeks old into 2 groups: group A was treated with 50 μg anti-CXCL13 monoclonal antibody through intraperitoneal injection once a day; group B was intraperitoneally administrated with 50 μg isotype Ig G once a day as a control group. 6 female C57 BL / 6 mice of 14 weeks old were used to set up group C as the normal control group. To evaluate renal function, freshly 24 h urine was collected at 14, 16, 18 and 20 w. The content of creatinine in serum and urine protein was tested by a Cr detection kit and a Radford protein quantification kit, respectively. Additionally,pathology damage of kidney was investigated by using hematoxylin and eosin staining. The results indicate proteinuria content in each group increased with time, but the increase rate over time in the group A was reduced. As compared with the group B, proteinuria was decreased by anti-CXCL13 monoclonal antibodies at 18 week(P<0.05). Similarly, the level of serum creatinine in group A was also reduced significantly when compared with the group B(P<0.01). According to HE staining, kidney damage degree of MRL/lpr mice in group A was lighter than the control group, with less inflammatory cell infiltration, glomerular mesenteric hyperplasia and glomerular mesangium matrix expansion. Taken together, anti-CXCL13 monoclonal antibody treatment can alleviate kidney damage in MRL/lpr mice.2. Influence of anti-CXCL13 monoclonal antibody treatment on LN in MRL/lpr miceThe grouping and dosing method are as above.In the present study, MRL/lpr mice were treated with anti-CXCL13 monoclonal antibodies and isotype Ig G, respectively. Then, immune complex deposition in glomeruli was investigated by using immunofluorescence. ELISA was employed to examine the level of anti-ds DNA in serum as well as content of inflammatory cytokines in renal tissues.By comparison with group B, Ig G and C3 immune complex deposition in MRL/LPR mice in group A was significantly reduced, and immune fluorescence intensity score was also reduced(P<0.01). Futhermore, level of anti-ds DNA antibodies in serum as well as inflammatory factors such as expression of IL-33, IL-1β, IL-6 and IL-17 significantly decreased( P<0.01). Taken together, anti-CXCL13 monoclonal antibody therapy may reduce kidney damage of MRL/LPR mice by reducing the level of the anti-ds DNA antibodies in serum, immune complex deposition in the kidney tissues and inflammatory factors in kidney tissues.3. Survival and Thl7/Treg ratio of MRL/LPR mice after treatment with anti-CXCL13 monoclonal antibodyThe grouping and dosing method are as above.In the present study, MRL/lpr mice were treated with CXCL13 antibodies and isotype Ig G, respectively. Th17/Treg ratio in the spleens was determined by using flow cytometry. Survival of MRL/lpr mice in vitro was also observed and recorded.Results shown the proportion of Th17 cells was decreased and that of Treg cells was increased in group A compared with group B, and as a result, the blocking of CXCL13 significantly reduced the Th17/Treg ratio in MRL/lpr mice(P<0.01). Vitro observation also found anti-CXCL13 monoclonal antibody improved survival of MRL/lpr mice(P<0.01). Taken together, anti-CXCL13 monoclonal antibody reduces Th17/Treg ratio and improves survival of MRL/lpr mice.The innovation of this study is as follows. Firstly, this study uses anti-CXCL13 monoclonal antibodies into the treatment of MRL/LPR mice and observes its therapeutic effect for the first time. Secondly, this study demonstrates that therapy of anti-CXCL13 monoclonal antibody can reduce urine protein and serum creatinine levels in MRL/LPR mice reduce the renal pathological damage degree for the first time. Also, treatment with CXCL13 monoclonal antibody can reduce the level of the anti ds DNA antibodies in serum, immune complex deposition in the kidney tissues and inflammatory factors in kidney tissues. Additionally, anti-CXCL13 monoclonal antibody reduces Th17/Treg ratio and improves survival of MRL/lpr mice. Thirdly, This study verified by animal experiments that anti-CXCL13 monoclonal antibodies is effective to lupus treatment and its mechanism may be related to the decrease of immune complex deposition. Anti-ds DNA antibodies, inflammatory factor levels and Thl7/Treg cells ratio. Anti-CXCL13 monoclonal antibody can be used as an effective therapeutic target in treatment of lupus nephritis of MRL/lpr mice, which has potential clinical utility value and provides a possible new way to the clinical treatment of LN. |
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