OBJECTIVE: To explore the construction of a canine model of vascularized allogeneic spinal cord transplantation (vASCT) and preliminarily evaluate its therapeutic efficacy for spinal cord injury (SCI). METHODS: Sixteen female Beagle dogs aged 8-12 months were randomly selected, with 8 dogs serving as donors for the harvesting of spinal cord tissue with a vascular pedicle [dorsal intercostal artery (DIA) at the T₁₀ level and accompanying vein]. The remaining 8 dogs underwent a 1.5-cm-length spinal cord defect at the T₁₀ level, followed by transplantation of the donor spinal cord tissue for repair. Polyethylene glycol (PEG) was applied to both ends to spinal cord graft; then, using a random number table method, the dogs were divided into an experimental group (n=4) and a control group (n=4). The experimental group received immunosuppressive intervention with oral tacrolimus [0.1 mg/(kg∙d)] postoperatively, while the control group received no treatment. The operation time and ischemia-reperfusion time of two groups were recorded. The recovery of hind limb function was estimated by Olby score within 2 months after operation; the motor evoked potentials (MEP) was measured through neuroelectrophysiological examination, and the spinal cord integrity was observed through MRI. RESULTS: There was no significant difference in the operation time and ischemia-reperfusion time between the two groups (P>0.05). All dogs survived until the completion of the experiment. Within 2 months after operation, all dogs in the control group failed to regain the movement function of hind limbs, and Olby scores were all 0. In the experimental group, the movement and weight-bearing, as well as walking abilities of the hind limbs gradually recovered, and the Olby scores also showed a gradually increasing trend. There was a significant difference between the two groups from 3 to 8 weeks after operation (P<0.05). Neuroelectrophysiological examination indicated that the electrical signals of the experimental group passed through the transplanted area, and the latency was shortened compared to that at 1 month after operation (P<0.05), showing continuous improvement, but the amplitude did not show significant improvement (P>0.05). The control group was unable to detect any MEP changes after operation. MRI examination showed that the transplanted spinal cord in the experimental group survived and had good continuity with normal spinal cord tissue, while no relevant change was observed in the control group. CONCLUSION The vASCT model of dogs was successfully constructed. This surgical procedure can restore the continuity of the spinal cord. The combination of tacrolimus anti-immunity is a key factor for the success of transplantation.
Animals ; Dogs ; Female ; Spinal Cord/blood supply* ; Spinal Cord Injuries/surgery* ; Transplantation, Homologous ; Disease Models, Animal ; Recovery of Function ; Plastic Surgery Procedures/methods* ; Tacrolimus ; Immunosuppressive Agents

Objective To investigate the value of repeat renal biopsy in the treatment and prognosis of nephrotic syndrome(NS)and acute kidney injury(AKI)following immunosuppressive therapy in patients with lupus nephritis(LN). Methods A retrospective analysis was conducted for the clinicopathological data and follow-up records of LN patients undergoing repeat renal biopsy at Peking Union Medical College Hospital from January 1,2009 to December 31,2021. Results A total of 76 patients(55 females,72.4%)were included in this study,with the mean age at the first biopsy being(29.0±10.4)years,the median inter-biopsy interval of 4.0(2.0,7.0) years,and the median total follow-up duration of 7.5(5.0,13.8)years.Pathological transformation occurred in 46(60.5%)patients,and 2 patients had comorbid diabetic nephropathy.At repeat renal biopsy,50(65.8%) patients presented NS.These patients demonstrated lower estimated glomerular filtration rate(eGFR)(P<0.001),higher chronicity index(CI)(P=0.029),and higher complement C3(P<0.001)and C4(P<0.001)levels than those with NS at the first renal biopsy(n=50).Among the 28(36.8%) patients with AKI at repeat renal biopsy,8(28.6%)experienced acute exacerbation of chronic renal insufficiency.These patients exhibited higher serum creatinine level(P=0.002),C4 level(P=0.033),CI(P=0.042),and prevalence of thrombotic microangiopathy(P=0.046)than the patients showing AKI at the first renal biopsy(n=16),while the activity index(AI)showed no significant difference(P=0.051).Over 50% of NS and AKI patients underwent treatment modifications post-repeat renal biopsy,with clinical remission rates comparable to those after the first renal biopsy(both P>0.05).Elevated CI(≥5,P=0.001)and serum creatinine(≥140 μmol/L,P<0.001)at repeat renal biopsy were identified as independent risk factors for poor prognosis.The patients with AKI at repeat renal biopsy had higher incidence of endpoint events than the non-AKI patients(P=0.015).Neither AKI at the first renal biopsy nor NS at both biopsies had significant associations with prognosis. Conclusions Repeat renal biopsy reveals not only sustained high disease activity but also accelerates chronic progression in LN patients,which underscore its critical role in guiding the therapy for severe LN post-immunosuppression.AKI,CI≥5,and serum creatinine ≥140 μmol/L at repeat renal biopsy are strongly associated with poor prognosis.
Humans ; Lupus Nephritis/drug therapy* ; Female ; Retrospective Studies ; Adult ; Male ; Prognosis ; Biopsy ; Kidney/pathology* ; Acute Kidney Injury/pathology* ; Nephrotic Syndrome/pathology* ; Glomerular Filtration Rate ; Young Adult ; Immunosuppressive Agents/therapeutic use* ; Middle Aged

Humans ; Drug Monitoring/methods* ; Child ; Inflammatory Bowel Diseases/drug therapy* ; Immunosuppressive Agents/therapeutic use* ; China ; Biological Products/therapeutic use*

Mycophenolic acid (MPA), the active moiety of both mycophenolate mofetil (MMF) and enteric-coated mycophenolate sodium (EC-MPS), serves as a primary immunosuppressant for maintaining solid organ transplants. Therapeutic drug monitoring (TDM) enhances treatment outcomes through tailored approaches. This study aimed to develop an evidence-based guideline for MPA TDM, facilitating its rational application in clinical settings. The guideline plan was drawn from the Institute of Medicine and World Health Organization (WHO) guidelines. Using the Delphi method, clinical questions and outcome indicators were generated. Systematic reviews, Grading of Recommendations Assessment, Development, and Evaluation (GRADE) evidence quality evaluations, expert opinions, and patient values guided evidence-based suggestions for the guideline. External reviews further refined the recommendations. The guideline for the TDM of MPA (IPGRP-2020CN099) consists of four sections and 16 recommendations encompassing target populations, monitoring strategies, dosage regimens, and influencing factors. High-risk populations, timing of TDM, area under the curve (AUC) versus trough concentration (C₀), target concentration ranges, monitoring frequency, and analytical methods are addressed. Formulation-specific recommendations, initial dosage regimens, populations with unique considerations, pharmacokinetic-informed dosing, body weight factors, pharmacogenetics, and drug‍-‍drug interactions are covered. The evidence-based guideline offers a comprehensive recommendation for solid organ transplant recipients undergoing MPA therapy, promoting standardization of MPA TDM, and enhancing treatment efficacy and safety.
Mycophenolic Acid/administration & dosage* ; Drug Monitoring/methods* ; Humans ; Organ Transplantation ; Immunosuppressive Agents/administration & dosage* ; Delphi Technique

Extensive epigenetic reprogramming involves in memory CD8+ T-cell differentiation. The elaborate epigenetic rewiring underlying the heterogeneous functional states of CD8+ T cells remains hidden. Here, we profile single-cell chromatin accessibility and map enhancer-promoter interactomes to characterize the differentiation trajectory of memory CD8+ T cells. We reveal that under distinct epigenetic regulations, the early activated CD8+ T cells divergently originated for short-lived effector and memory precursor effector cells. We also uncover a defined epigenetic rewiring leading to the conversion from effector memory to central memory cells during memory formation. Additionally, we illustrate chromatin regulatory mechanisms underlying long-lasting versus transient transcription regulation during memory differentiation. Finally, we confirm the essential roles of Sox4 and Nrf2 in developing memory precursor effector and effector memory cells, respectively, and validate cell state-specific enhancers in regulating Il7r using CRISPR-Cas9. Our data pave the way for understanding the mechanism underlying epigenetic memory formation in CD8+ T-cell differentiation.
CD8-Positive T-Lymphocytes/metabolism* ; Cell Differentiation ; Chromatin/immunology* ; Animals ; Mice ; Immunologic Memory ; Epigenesis, Genetic ; SOXC Transcription Factors/immunology* ; NF-E2-Related Factor 2/immunology* ; Mice, Inbred C57BL ; Gene Regulatory Networks ; Enhancer Elements, Genetic

Objective To prepare mouse monoclonal antibodies against the ectodomain of E2 (E2ecto) glycoprotein of Western equine encephalitis virus (WEEV). Methods A prokaryotic expression plasmid pET-28a-WEEV E2ecto was constructed and transformed into BL21 (DE3) competent cells. E2ecto protein was expressed by IPTG induction and presented mainly as inclusion bodies. Then the purified E2ecto protein was prepared by denaturation, renaturation and ultrafiltration. BALB/c mice were immunized with the formulated E2ecto protein using QuickAntibody-Mouse5W as an adjuvant via intramuscular route, boosted once at an interval of 21 days. At 35 days post-immunization, mice with antibody titer above 1×10⁴ were inoculated with E2ecto intraperitoneally, and spleen cells were fused with SP2/0 cells three days later. Hybridoma cells secreting specific monoclonal antibodies were screened by the limited dilution method, and ascites were prepared after intraperitoneal inoculation of hybridoma cells. The subtypes and titers of the antibodies in ascites were assayed by ELISA. The biological activity of the mAb was identified by immunofluorescence assay(IFA) on BHK-21 cells which were transfected with eukaryotic expression plasmid pCAGGS-WEEV-CE3E2E1. The specificity of the antibodies were evaluated with E2ecto proteins from EEEV and VEEV. Results Purified WEEV E2ecto protein was successfully expressed and obtained. Four monoclonal antibodies, 3G6G10, 3D7G2, 3B9E8 and 3D5B7, were prepared, and their subtypes were IgG2c(κ), IgM(κ), IgM(κ) and IgG1(κ), respectively. The titers of ascites antibodies 3G6G10, 3B9E8 and 3D7G2 were 10⁵, and 3D5B7 reached 10⁷. None of the four antibody strains cross-reacted with other encephalitis alphavirus such as VEEV and EEEV. Conclusion Four strains of mouse mAb specifically binding WEEV E2ecto are successfully prepared.
Horses ; Animals ; Mice ; Encephalitis Virus, Western Equine ; Ascites ; Immunosuppressive Agents ; Antibodies, Monoclonal ; Immunoglobulin M

OBJECTIVE: The present study aimed to evaluate the immunogenicity of BA.2 variant receptor binding domain (RBD) recombinant protein formulated with CpG 1826 plus alum dual adjuvant. METHODS: The BA.2 variant RBD (residues 308-548) fusing TT-P ₂ epitope was obtained from prokaryotic expression system, purification technology and dialysis renaturation, which was designated as Sot protein. The soluble Sot protein formulated with CpG 1826 plus alum dual adjuvant was designated as Sot/CA subunit vaccine and then the BALB/c mice were intramuscularly administrated with two doses of the Sot/CA subunit vaccine at 14-day interval (day 0 and 14). On day 28, the number of effector T lymphocytes secreting IFN-γ and IL-4 in mice spleen were determined by enzyme-linked immunospot (ELISpot) assay. The serum IgG, IgG1 and IgG2a antibodies were examined by enzyme-linked immunosorbent assay (ELISA). In addition, the level of neutralizing antibodies (NAbs) induced by Sot/CA subunit vaccine was also evaluated by the microneutralization assay. RESULTS: The high-purity soluble Sot protein with antigenicity was successfully obtained by the prokaryotic expression, protein purification and dialysis renaturation. The Sot/CA subunit vaccine induced a high level of IgG antibodies and NAbs, which were of cross-neutralizing activity against SARS-CoV-2 BA.2 and XBB.1.5 variants. Meanwhile, Sot/CA subunit vaccine also induced a high level of effector T lymphocytes secreting IFN-γ (635.00 ± 17.62) and IL-4 (279.20 ± 13.10), respectively. Combined with a decreased IgG1/IgG2a ratio in the serum, which indicating Sot/CA subunit vaccine induced a Th1-type predominant immune response. CONCLUSION The Sot protein formulated with CpG 1826 plus alum dual adjuvant showed that the excellent cellular and humoral immunogenicity, which provided a scientific basis for the development of BA.2 variant subunit vaccines and references for the adjuvant application of subunit vaccines.
Animals ; COVID-19 Vaccines/immunology* ; Alum Compounds/pharmacology* ; Mice, Inbred BALB C ; Vaccines, Subunit/immunology* ; Mice ; SARS-CoV-2/immunology* ; Oligodeoxyribonucleotides/administration & dosage* ; Female ; Adjuvants, Immunologic ; COVID-19/immunology* ; Antibodies, Viral/blood* ; Immunogenicity, Vaccine ; Spike Glycoprotein, Coronavirus/immunology* ; Antibodies, Neutralizing/blood* ; Adjuvants, Vaccine ; Immunoglobulin G/blood*

Vaccination is an effective method for preventing influenza, and adjuvants can enhance the immune response intensity and persistence of influenza vaccines. However, there are currently shortcomings in clinical adjuvant approvals, ineffectiveness against weak antigens, and a tendency to cause headaches. Therefore, the development of safe and effective novel adjuvants for influenza vaccines is particularly important to enhance vaccine immunogenicity and safety. Given the wide range of sources, high safety, and biodegradability of traditional Chinese medicine(TCM), some studies have described it as a vaccine adjuvant. This article reviewed the current status and challenges of influenza vaccine adjuvants, summarized the types of TCM adjuvants, the safety and immunomodulatory effects of natural active ingredients from TCM combined with influenza vaccines, the role of TCM adjuvants in antigen storage, antigen presentation capability, immune cells and cytokines, and immune responses, and analyzed the advantages and disadvantages of TCM adjuvants compared with small molecule adjuvants, with the aim of promoting the clinical development and commercialization of TCM adjuvants for influenza vaccines.
Humans ; Influenza Vaccines ; Adjuvants, Immunologic/pharmacology* ; Medicine, Chinese Traditional ; Influenza, Human/prevention & control* ; Adjuvants, Pharmaceutic

OBJECTIVES: To investigate the clinical effect of different immunosuppressive treatment regimens in children with ocular myasthenia gravis (OMG). METHODS: A retrospective analysis was conducted on 130 children with OMG who were treated in the Department of Neurology, Jiangxi Children's Hospital, from February 2018 to February 2023. According to the treatment regimen, they were divided into four groups: glucocorticoid (GC) group (n=29), mycophenolate mofetil (MMF) group (GC+MMF; n=33), methotrexate (MTX) group (GC+MTX; n=30), and tacrolimus (FK506) group (GC+FK506; n=38). Treatment outcomes and adverse reactions were compared among the groups. RESULTS: After 3 months of treatment, the FK506 group had significantly lower scores of Myasthenia Gravis Quantitative Scale and Myasthenia Gravis-Specific Activities of Daily Living than the other three groups (P<0.05). After 3 months of treatment, the FK506 group had a significantly lower dose of prednisone than the GC group, and after 6 and 9 months of treatment, the MMF, MTX, and FK506 groups had a significantly lower dose of prednisone than the GC group (P<0.05). After 12 months of treatment, the MMF, MTX, and FK506 groups had a significantly lower incidence rate of GC-related adverse reactions than the GC group (P<0.05). CONCLUSIONS For children with OMG, the addition of various immunosuppressants can reduce the dosage of GC and adverse reactions. Among them, FK506 shows superior efficacy compared to other immunosuppressants in the early treatment of OMG.
Humans ; Child ; Prednisone/adverse effects* ; Tacrolimus/adverse effects* ; Retrospective Studies ; Activities of Daily Living ; Immunosuppressive Agents/adverse effects* ; Myasthenia Gravis/drug therapy* ; Glucocorticoids/therapeutic use* ; Mycophenolic Acid/adverse effects*

OBJECTIVES: To investigate the difference in the therapeutic effect of mycophenolate mofetil (MMF) or cyclophosphamide (CTX) in children with Henoch-Schönlein purpura nephritis (HSPN) of different age groups. METHODS: A retrospective analysis was conducted on the clinical data of 135 children with HSPN who were treated with MMF or CTX in the Department of Nephrology, Children's Hospital Affiliated to Capital Institute of Pediatrics, from October 2018 to October 2020. According to the immunosuppressant used, they were divided into two groups: MMF group and CTX group, and according to the age, each group was further divided into two subgroups: ≤12 years and >12 years, producing four groups, i.e, the ≤12 years MMF subgroup (n=30), the >12 years MMF subgroup (n=15), the ≤12 years CTX subgroup (n=71), and the >12 years CTX subgroup (n=19). All children were followed up for at least 12 months, and the above groups were compared in terms of clinical outcomes and the incidence rate of adverse reactions. RESULTS: There was no significant difference in the complete response rate between the MMF group and the CTX group after 3, 6, and 12 months of treatment (P>0.05). There were no significant difference in the complete response rate and the incidence rate of adverse reactions between the >12 years MMF subgroup and the ≤12 years MMF subgroup at 3, 6, and 12 months of treatment (P>0.05). The >12 years CTX subgroup had a significantly lower complete response rate than the ≤12 years CTX subgroup at 6 and 12 months of treatment (P<0.05). The >12 years CTX subgroup had a significantly higher incidence rate of adverse reactions than the >12 years MMF subgroup (P<0.05). CONCLUSIONS The efficacy and adverse reactions of MMF are not associated with age, but the efficacy of CTX is affected by age, with a higher incidence rate of adverse reactions. CTX should be selected with caution for children with HSPN aged >12 years.
Child ; Humans ; Mycophenolic Acid/adverse effects* ; IgA Vasculitis/drug therapy* ; Retrospective Studies ; Cyclophosphamide/adverse effects* ; Immunosuppressive Agents/adverse effects* ; Vasculitis/drug therapy* ; Nephritis/complications*