IgG subclass profile among anti‐Glutathione S‐transferase T1 antibodies in post‐transplant de novo immune hepatitis

Although the pathogenic pathways leading to de novo immune hepatitis (IH) are not completely understood, we have shown strong evidences of an antidonor response against Glutathione S‐transferase T1 (GSTT1), an antigen exclusively expressed in the donor liver. The first sign of this process is the production of GSTT1 antibodies that, in 25% of the cases, will precede de novo IH. Because the presence of the antibodies is not sufficient to trigger the disease, we aimed to study GSTT1 IgG subclasses in a group of 18 liver transplant patients, 12 that developed de novo IH and 6 that remained free of disease. Surprisingly, the predominant subclasses were IgG1‐_GSTT1 and IgG4‐_GSTT1. The presence of IgG4‐expressing plasma cells was also investigated in 10 available liver biopsies. Six biopsies coinciding with diagnosis showed a mean value of 32.8 IgG4+ plasma cells/hpf vs. 5.55 in patients without the disease. We have not found a distinctive GSTT1‐IgG profile in patients with de novo IH, but the ratio IgG1‐_GSTT1/IgG4‐_GSTT1 in samples from close to the time of diagnosis seemed to be important. The novel finding of abundant IgG4‐_GSTT1 in liver transplantation is intriguing, but their possible role in pathogenesis of de novo IH remains unknown.     

Human pregnancy and generation of anti‐angiotensin receptor and anti‐perlecan antibodies

Non‐HLA antibodies against the angiotensin II type 1 receptor (AT₁R) and the C‐terminal fragment of perlecan (i.e., LG3) are associated with the development of renal allograft rejection. It is currently unknown how humans develop anti‐AT₁R or anti‐LG3 antibodies. The aim of this study was to investigate whether pregnancy—as a model of sensitization to polymorphic proteins—induces anti‐AT₁R and/or anti‐LG3 antibodies. We included 104 samples from women obtained after physiologic full‐term pregnancy and 80 samples from healthy nonsensitized controls (40 women and 40 men). Both anti‐AT₁R and anti‐LG3 antibody levels were lower in pregnancy samples than in controls (both P < 0.05). By multivariate analysis, male gender was an independent predictor for high anti‐AT₁R antibody levels (OR 3.66, P = 0.04) and pregnancy was predictive for low anti‐LG3 antibody levels (OR 6.53, P = 0.0001). There was no correlation of anti‐AT₁R with anti‐LG3 antibody levels, either in the pregnancy or in the control samples (r² ≤ 0.03, P ≥ 0.26). In conclusion, physiologic full‐term pregnancy does not induce anti‐AT₁R or anti‐LG3 antibodies and may even lower their levels. Therefore, anti‐AT₁R and anti‐LG3 antibodies are likely not caused by allosensitization. The lack of correlation of anti‐AT₁R with anti‐LG3 antibodies suggests different mechanisms of generation, which remain to be elucidated.     

Novel quinolone CHM‐1 induces apoptosis and inhibits metastasis in a human osterogenic sarcoma cell line

Novel 2‐phenyl‐4‐quinolone compounds have potent cytotoxic effects on different human cancer cell lines. In this study, we examined anticancer activity and mechanisms of 20‐fluoro‐6,7‐methylenedioxy‐2‐phenyl‐4‐quinolone (CHM‐1) in human osterogenic sarcoma U‐2 OS cells. CHM‐1‐induced apoptosis was determined by flow cytometric analysis, DAPI staining, Comet assay, and caspase inhibitors. CHM‐1‐inhibited cell migration and invasion was assessed by a wound healing assay, gelatin zymography, and a Transwell assay. The mechanisms of CHM‐1 effects on apoptosis and metastasis signaling pathways were studied using Western blotting and gene expression. CHM‐1 induced G2/M arrest and apoptosis at an IC₅₀ (3 µM) in U‐2 OS cells and caspase‐3, ‐8, and ‐9 were activated. Caspase inhibitors increased cell viability after exposure to CHM‐1. CHM‐1‐induced apoptosis was associated with enhanced ROS generation, DNA damage, decreased ΔΨ_m levels, and promotion of mitochondrial cytochrome c release. CHM‐1 stimulated mRNA expression of caspase‐3, ‐8, and ‐9, AIF, and Endo G. In addition, CHM‐1 inhibited cell metastasis at a low concentration (<3 µM). CHM‐1 inhibited the cell metastasis through the inhibition of MMP‐2, ‐7, and ‐9. CHM‐1 also decreased the levels of MAPK signaling pathways before leading to the inhibition of MMPs. In summary, CHM‐1 is a potent inducer of apoptosis, which plays a role in the anticancer activity of CHM‐1. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:1637–1644, 2009     

Thyroid hormone‐mediated growth and differentiation of growth plate chondrocytes involves IGF‐1 modulation of β‐catenin signaling

Thyroid hormone regulates terminal differentiation of growth plate chondrocytes in part through modulation of the Wnt/β‐catenin signaling pathway. Insulin‐like growth factor 1 (IGF‐1) has been described as a stabilizer of β‐catenin, and thyroid hormone is a known stimulator of IGF‐1 receptor expression. The purpose of this study was to test the hypothesis that IGF‐1 signaling is involved in the interaction between the thyroid hormone and the Wnt/β‐catenin signaling pathways in regulating growth plate chondrocyte proliferation and differentiation. The results show that IGF‐1 and the IGF‐ receptor (IGF1R) stimulate Wnt‐4 expression and β‐catenin activation in growth plate chondrocytes. The positive effects of IGF‐1/IGF1R on chondrocyte proliferation and terminal differentiation are partially inhibited by the Wnt antagonists sFRP3 and Dkk1. T₃ activates IGF‐1/IGF1R signaling and IGF‐1‐dependent PI3K/Akt/GSK‐3β signaling in growth plate chondrocytes undergoing proliferation and differentiation to prehypertrophy. T₃‐mediated Wnt‐4 expression, β‐catenin activation, cell proliferation, and terminal differentiation of growth plate chondrocytes are partially prevented by the IGF1R inhibitor picropodophyllin as well as by the PI3K/Akt signaling inhibitors LY294002 and Akti1/2. These data indicate that the interactions between thyroid hormone and β‐catenin signaling in regulating growth plate chondrocyte proliferation and terminal differentiation are modulated by IGF‐1/IGF1R signaling through both the Wnt and PI3K/Akt signaling pathways. While chondrocyte proliferation may be triggered by the IGF‐1/IGF1R‐mediated PI3K/Akt/GSK3β pathway, cell hypertrophy is likely due to activation of Wnt/β‐catenin signaling, which is at least in part initiated by IGF‐1 signaling or the IGF‐1‐activated PI3K/Akt signaling pathway. © 2010 American Society for Bone and Mineral Research     

Detection of New Delhi Metallo‐beta‐Lactamase and Extended‐Spectrum beta‐Lactamase Genes in Escherichia coli Isolated from Mastitic Milk Samples

In this study, eight Escherichia coli isolates were obtained from milk samples of dairy cattle suffering from clinical/subclinical mastitis. Isolates were characterized for antimicrobial resistance traits and virulence genes. Results revealed that one isolate was harbouring New Delhi metallo‐beta‐lactamase gene (bla_NDM). Cloning and sequencing of the PCR amplicon confirmed the identity of the gene (GenBank accession no. KC769583 ) having 100% homology with bla_NDM‐5 (GenBank accession no. JN104597.1 ), and this isolate was susceptible to colistin, chloramphenicol and tetracycline only. Moreover, another isolate carried extended‐spectrum beta‐lactamase (ESBL) gene – bla_CTX‐M, and all isolates possessed bla_TEM gene. Of the eight isolates, only one isolate was positive for shiga toxin gene (stx2), and none were harbouring stx1 gene. Occurrence of New Delhi metallo‐beta‐lactamase (bla_NDM) in one E. coli isolate and ESBL genes in other isolates poses a potential threat to human health following possible entry and spread through food chain.     

Combined treatment with a β3‐adrenergic receptor agonist and a muscarinic receptor antagonist inhibits detrusor overactivity induced by cold stress in spontaneously hypertensive rats

Aims

This study determined if combined treatment with the muscarinic receptor (MR) antagonist solifenacin and the β₃‐adrenergic receptor (AR) agonist mirabegron could inhibit detrusor overactivity induced by cold stress in spontaneously hypertensive rats (SHRs).

Methods

Thirty‐two female 10‐week‐old SHRs were fed an 8% NaCl‐supplemented diet for 4 weeks. Cystometric measurements of the unanesthetized, unrestricted rats were performed at room temperature (RT, 27 ± 2°C) for 20 min. The rats were then intravenously administered vehicle, 0.1 mg/kg solifenacin alone, 0.1 mg/kg mirabegron alone, or the combination of 0.1 mg/kg mirabegron and 0.1 mg/kg solifenacin (n = 8 each group). Five minutes later, the treated rats were exposed to low temperature (LT, 4 ± 2°C) for 40 min. Finally, the rats were returned to RT. After the cystometric investigations, the β₃‐ARs and M₃‐MRs expressed within the urinary bladders were analyzed.

Results

Just after transfer from RT to LT, vehicle‐, solifenacin‐, and mirabegron‐treated SHRs exhibited detrusor overactivity that significantly decreased voiding interval and bladder capacity. However, treatment with the combination of solifenacin and mirabegron partially inhibited the cold stress‐induced detrusor overactivity patterns. The decreases of voiding interval and bladder capacity in the combination‐treated rats were significantly inhibited compared to other groups. Within the urinary bladders, there were no differences between expression levels of M₃‐MR and β₃‐AR mRNA. The tissue distribution of M₃‐MRs was similar to that of the β₃‐ARs.

Conclusions

This study suggested that the combination of solifenacin and mirabegron act synergistically to inhibit the cold stress‐induced detrusor overactivity in SHRs. Neurourol. Urodynam. 36:1026–1033, 2017. © 2016 The Authors. Neurourology and Urodynamics Published by Wiley Periodicals, Inc.     

Improved Screening Test for Idiopathic Infantile Hypercalcemia Confirms Residual Levels of Serum 24,25‐(OH)2D3 in Affected Patients

CYP24A1 mutations are now accepted as a cause of idiopathic infantile hypercalcemia (IIH). A rapid liquid‐chromatography tandem mass spectrometry (LC‐MS/MS)‐based blood test enabling measurement of the 25‐OH‐D₃:24,25‐(OH)₂D₃ ratio (R) can identify IIH patients on the basis of reduced C24‐hydroxylation of 25‐OH‐D₃ by CYP24A1 in vivo. Although values of this ratio are significantly elevated in IIH, somewhat surprisingly, serum 24,25‐(OH)₂D₃ remains detectable. The current study explores possible explanations for this including: residual CYP24A1 enzyme activity in individuals with certain CYP24A1 genotypes, expression of alternative C24‐hydroxylases, and the possibility of isobaric contamination of the 24,25‐(OH)₂D₃ peak on LC‐MS/MS. We employed an extended 20‐min run time on LC‐MS/MS to study serum vitamin D metabolites in patients with IIH due to mutations of CYP24A1 or SLC34A1; in unaffected heterozygotes and dialysis patients; in patients with vitamin D deficiency; as well as in normal subjects exhibiting a broad range of 25‐OH‐D levels. We identified 25,26‐(OH)₂D₃ as a contaminant of the 24,25‐(OH)₂D₃ peak. In normals, the concentration of 24,25‐(OH)₂D₃ greatly exceeds 25,26‐(OH)₂D₃; however, 25,26‐(OH)₂D₃ becomes more significant in IIH with CYP24A1 mutations and in dialysis patients, where 24,25‐(OH)₂D₃ levels are low when CYP24A1 function is compromised. Mean R in 30 IIH‐CYP24A1 patients was 700 (range, 166 to 2168; cutoff = 140) as compared with 31 in 163 controls. Furthermore, patients possessing CYP24A1 L409S alleles exhibited higher 24,25‐(OH)₂D₃ levels and lower R (mean R = 268; n = 8) than patients with other mutations. We conclude that a chromatographic approach which resolves 24,25‐(OH)₂D₃ from 25,26‐(OH)₂D₃ produces a more accurate R that can be used to differentiate pathological states where CYP24A1 activity is altered. The origin of the residual serum 24,25‐(OH)₂D₃ in IIH patients appears to be multifactorial. © 2017 American Society for Bone and Mineral Research.     

Enhancement of hepatic 4‐hydroxylation of 25‐hydroxyvitamin D3 through CYP3A4 induction in vitro and in vivo: Implications for drug‐induced osteomalacia

Long‐term therapy with certain drugs, especially cytochrome P450 (P450; CYP)‐inducing agents, confers an increased risk of osteomalacia that is attributed to vitamin D deficiency. Human CYP24A1, CYP3A4, and CYP27B1 catalyze the inactivation and activation of vitamin D and have been implicated in the adverse drug response. In this study, the inducibility of these enzymes and monohydroxylation of 25‐hydroxyvitamin D₃ (25OHD₃) were evaluated after exposure to P450‐inducing drugs. With human hepatocytes, treatment with phenobarbital, hyperforin, carbamazepine, and rifampin significantly increased the levels of CYP3A4, but not CYP24A1 or CYP27B1 mRNA. In addition, rifampin pretreatment resulted in an 8‐fold increase in formation of the major metabolite of 25OHD₃, 4β,25(OH)₂D₃. This inductive effect was blocked by the addition of 6′,7′‐dihydroxybergamottin, a selective CYP3A4 inhibitor. With human renal proximal tubular HK‐2 cells, treatment with the same inducers did not alter CYP3A4, CYP24A1, or CYP27B1 expression. 24R,25(OH)₂D₃ was the predominant monohydroxy metabolite produced from 25OHD₃, but its formation was unaffected by the inducers. With healthy volunteers, the mean plasma concentration of 4β,25(OH)₂D₃ was increased 60% (p < 0.01) after short‐term rifampin administration. This was accompanied by a statistically significant reduction in plasma 1α,25(OH)₂D₃ (?10%; p = 0.03), and a nonsignificant change in 24R,25(OH)₂D₃ (?8%; p = 0.09) levels. Further analysis revealed a negative correlation between the increase in 4β,25(OH)₂D₃ and decrease in 1α,25(OH)₂D₃ levels. Examination of the plasma monohydroxy metabolite/25OHD₃ ratios indicated selective induction of the CYP3A4‐dependent 4β‐hydroxylation pathway of 25OHD₃ elimination. These results suggest that induction of hepatic CYP3A4 may be important in the etiology of drug‐induced osteomalacia. © 2013 American Society for Bone and Mineral Research.     

Elevated serum IGF‐1 levels synergize PTH action on the skeleton only when the tissue IGF‐1 axis is intact

There is growing evidence that insulin‐like growth factor 1 (IGF‐1) and parathyroid hormone (PTH) have synergistic actions on bone and that part of the anabolic effects of PTH is mediated by local production of IGF‐1. In this study we analyzed the skeletal response to PTH in mouse models with manipulated endocrine or autocrine/paracrine IGF‐1. We used mice carrying a hepatic IGF‐1 transgene (HIT), which results in a threefold increase in serum IGF‐1 levels and normal tissue IGF‐1 expression, and Igf1 null mice with blunted IGF‐1 expression in tissues but threefold increases in serum IGF‐1 levels (KO‐HIT). Evaluation of skeletal growth showed that elevations in serum IGF‐1 in mice with Igf1 gene ablation in all tissues except the liver (KO‐HIT) resulted in a restoration of skeletal morphology and mechanical properties by adulthood. Intermittent PTH treatment of adult HIT mice resulted in increases in serum osteocalcin levels, femoral total cross‐sectional area, cortical bone area and cortical bone thickness, as well as bone mechanical properties. We found that the skeletal response of HIT mice to PTH was significantly higher than that of control mice, suggesting synergy between IGF‐1 and PTH on bone. In sharp contrast, although PTH‐treated KO‐HIT mice demonstrated an anabolic response in cortical and trabecular bone compartments compared with vehicle‐treated KO‐HIT mice, their response was identical to that of PTH‐treated control mice. We conclude that (1) in the presence of elevated serum IGF‐1 levels, PTH can exert an anabolic response in bone even in the total absence of tissue IGF‐1, and (2) elevations in serum IGF‐1 levels synergize PTH action on bone only if the tissue IGF‐1 axis is intact. Thus enhancement of PTH anabolic actions depends on tissue IGF‐1. © 2010 American Society for Bone and Mineral Research.     

Compression‐induced HIF‐1 enhances thrombosis and PAI‐1 expression in mouse skin

Pressure ulcers result from tissue hypoxia caused by external forces. Thrombosis due to external forces is considered important, and hypoxia inducible factor‐1 (HIF‐1) is a master regulator of pressure ulcer development. To date, however, their causal relationship has not been determined. This study therefore investigated the mutual relationship between thrombosis and HIF‐1 activation in compressed mouse skin, based on a hypothesis that HIF‐1 regulation by plasminogen activator inhibitor‐1 (PAI‐1) enhances thrombosis. Compression of mouse skin significantly increased the numbers of thrombi and HIF‐1α‐positive cells compared with control skin. A thrombosis inhibitor significantly reduced the numbers of HIF‐1α‐positive cells and an HIF‐1 inhibitor significantly inhibited thrombosis in compressed skin tissue, suggesting a mutual relationship between thrombosis and HIF‐1 activation. Compression of mouse skin also enhanced the level of Pai‐1 messenger RNA expression, but this increase was significantly reduced by treatment with an HIF‐1 inhibitor, whereas a thrombosis inhibitor had no effect. These results suggested the involvement of PAI‐1 in HIF‐1‐enhanced thrombosis and that an additional factor participates in regulating Pai‐1 expression in compressed skin. These findings may suggest new strategies in pressure ulcer management.     

The impact of donor body mass index on outcomes after deceased kidney transplantation – a national population‐cohort study

The aim of this study was to determine the effect of donor body mass index (BMI) on deceased donor kidney transplant outcomes. Data were collected from the UK Transplant Registry for all deceased donor kidney transplant recipients between January 2003 and January 2015. Univariable and multivariable analyses were undertaken to assess the impact of donor BMI on a range of outcomes. Donor BMI (kg/m²) was stratified as <18.5 (n = 380), 18.5–25.0 (n = 6890), 25.1–30.0 (n = 6669), 30.1–35.0 (n = 2503) and >35.0 (n = 1148). The prevalence of delayed graft function increased significantly with donor BMI (P < 0.001), with an adjusted odds ratio of 1.38 (95% CI: 1.16–1.63) for the >35.0 vs. 18.5–25.0 groups. However, there was no significant association between donor BMI and 12‐month creatinine (P = 0.550), or patient (P = 0.109) or graft (P = 0.590) survival. In overweight patients, increasing donor BMI was associated with a significant increase in warm ischaemia time and functional warm ischaemia time, by an average of 4.6% (P = 0.043) and 5.2% (P = 0.013) per 10.0 kg/m². However, rising warm ischaemic time and functional warm ischaemic time was not significantly associated with delayed graft function, 12‐month creatinine levels, graft loss or patient death. In this population cohort study, we identified no significant association between donor BMI and long‐term clinical outcomes in deceased donor kidney transplantation.     

Endothelin‐1 Acutely Reduces the Permeability of Visceral Sheep Peritoneum In Vitro Through Both Endothelin‐A and Endothelin‐B Receptors

Mesothelium is an important part of the peritoneal barrier for water and ion transport, essential for effective peritoneal dialysis (PD). Peritoneal fibrosis has been associated with PD treatment failure. Endothelin‐1 (ET‐1) is a potent vasoactive peptide, involved in pathologic fibrotic processes. Its action is mediated mainly by endothelin type A (ET_A) and type B (ET_B) receptors. The aim of this study was to investigate, by Ussing chamber experiments, the effect of ET‐1 on the transmesothelial electrical resistance (R_TM) of the isolated visceral sheep peritoneum. Intact sheets of visceral peritoneum were obtained from 40 adult sheep and mounted in Ussing‐type chambers. ET‐1 (10^?7 M), BQ‐123 (ET_A receptor antagonist; 10^?6 M), BQ‐788 (ET_B receptor antagonist; 10^?6 M), and their combinations were added on the apical and the basolateral side of the peritoneum. R_TM was measured before and serially after addition of the substances, and changes were registered as percentage (ΔR_TM %). R_TM increased within 1 min after addition of ET‐1 apically (ΔR_TM 65.03 ± 15.87%; P < 0.05) or basolaterally (ΔR_TM 85.5 ± 20.86%; P < 0.05). BQ‐123 and BQ‐788 and their combination significantly reduced (P < 0.05) the effect of ET‐1 to a similar degree in all cases. These results clearly indicate that ET‐1 reduces ionic permeability of the visceral sheep peritoneum in vitro. Additionally, it is obvious that this inhibitory effect is mediated through both ET_A and ET_B receptors.     

Enhanced effect of human mesenchymal stem cells expressing human TNF‐αR‐Fc and HO‐1 gene on porcine islet xenotransplantation in humanized mice

Background

Porcine islet xenotransplantation is considered an attractive alternative treatment for type 1 diabetes mellitus. However, it is largely limited because of initial rejection due to Instant Blood‐Mediated Inflammatory Reaction (IBMIR), oxidative stress, and inflammatory responses. Recently, soluble tumor necrosis factor‐ɑ receptor type I (sTNF‐αR) and heme oxygenase (HO)‐1 genes (HO‐1/sTNF‐αR) have been shown to improve the viability and functionality of porcine islets after transplantation.

Methods

In this study, genetically modified mesenchymal stem cells (MSCs) expressing the HO‐1/sTNF‐αR genes (HO‐1/sTNF‐αR‐MSC) were developed using an adenoviral system, and porcine islet viability and function were confirmed by in vitro tests such as GSIS, AO/PI, and the ADP/ATP ratio after coculturing with HO‐1/sTNF‐αR‐MSCs. Subsequently, isolated porcine islets were transplanted underneath the kidney capsule of diabetic humanized mice without MSCs, with MSCs or with HO‐1/sTNF‐αR‐MSCs.

Results

According to the results, the HO‐1/sTNF‐αR‐MSC‐treated group exhibited improved survival of porcine islets and could reverse hyperglycemia more than porcine islets not treated with MSCs or islets cotransplanted with MSCs. Moreover, the HO‐1/sTNF‐αR‐MSC group maintained its morphological characteristics and the insulin secretion pattern of transplanted porcine islets similar to endogenous islets in immunocompetent humanized mice.

Conclusions

Our results suggest that HO‐1/sTNF‐αR‐MSCs are efficient tools for porcine islet xenotransplantation, and this study may provide basic information for pre‐clinical animal models and future clinical trials of porcine islet xenotransplantation.     

Nitric Oxide Ventilation of Rat Lungs from Non‐Heart‐Beating Donors Improves Posttransplant Function

Lungs from non‐heart‐beating donors (NHBDs) would enhance the donor pool. Ex vivo perfusion and ventilation of NHBD lungs allows functional assessment and treatment. Ventilation of rat NHBD lungs with nitric oxide (NO) during ischemia, ex vivo perfusion and after transplant reduced ischemia‐reperfusion injury (IRI) and improved lung function posttransplant. One hour after death, Sprague‐Dawley rats were ventilated for another hour with either 60% O2 or 60% O2/40 ppm NO. Lungs were then flushed with 20‐mL cold Perfadex, stored cold for 1 h, perfused in an ex vivo circuit with Steen solution and warmed to 37°C, ventilated 15 min, perfusion‐cooled to 20°C, then flushed with cold Perfadex and stored cold. The left lung was transplanted and ventilated separately. Recipients were sacrificed after 1 h. NO‐ventilation was associated with significantly reduced wet:dry weight ratio in the ex vivo circuit, better oxygenation, reduced pulmonary vascular resistance, increased lung tissue levels of cGMP, maintained endothelial NOS eNOS, and reduced increases in tumor necrosis factor alpha (TNF‐α) and inducible nitric oxide synthase (iNOS). NO‐ventilation had no effect on MAP kinases or NF‐κB activation. NO administration to NHBDs before and after lung retrieval may improve function of lungs from NHBDs.     

De novo donor‐specific anti‐HLA antibodies mediated rejection in liver‐transplant patients

The incidence and consequences of de novo donor‐specific anti‐HLA antibodies (DSAs) after liver transplantation (LT) are not well known. We investigated the incidence, risk factors, and complications associated with de novo DSAs in this setting. A total of 152 de novo liver‐transplant patients, without preformed anti‐HLA DSAs, were tested for anti‐HLA antibodies, with single‐antigen bead technology, before, at transplantation, at 1, 3, 6 and 12 months after transplantation, and thereafter annually and at each time they presented with increased liver‐enzyme levels until the last follow‐up, that is, 34 (1.5–77) months. Twenty‐one patients (14%) developed de novo DSAs. Of these, five patients had C1q‐binding DSAs (24%). Younger age, low exposure to calcineurin inhibitors, and noncompliance were predictive factors for de novo DSA formation. Nine of the 21 patients (43%) with de novo DSAs experienced an acute antibody‐mediated rejection (AMR). Positive C4d staining was more frequently observed in liver biopsies of patients with AMR (9/9 vs. 1/12, P < 0.0001). Eight patients received a B‐cell targeting therapy, and one patient received polyclonal antibodies. Only one patient required retransplantation. Patient‐ and graft‐survival rates did not differ between patients with and without DSAs. In conclusion, liver‐transplant patients with liver abnormalities should be screened for DSAs and AMR.     

Galectin‐1 Prolongs Survival of Mouse Liver Allografts From Flt3L‐Pretreated Donors

Liver allografts are spontaneously accepted across MHC barriers in mice. The mechanisms underlying this phenomenon remain poorly understood. Galectin‐1, an endogenous lectin expressed in lymphoid organs, plays a vital role in maintaining central and peripheral tolerance. This study was to investigate the role of galectin‐1 in spontaneous tolerance of liver allografts in mice, and to evaluate the therapeutic effects of galectin‐1 on liver allograft rejection induced by donor Flt3L pretreatment. Blockade of the galectin‐1 pathway via neutralizing antigalectin‐1 mAb did not affect survival of the liver allografts from B6 donors into C3H recipients. Administration of rGal‐1 significantly prolonged survival of liver allografts from Flt3L‐pretreated donors and ameliorated Flt3L‐triggered liver allograft rejection. This effect was associated with increased apoptosis of T cells in both allografts and spleens, decreased frequencies of Th1 and Th17 cells, decreased expression of Th1‐associated cytokines (IL‐12, IL‐2 and IFN‐γ), Th17‐associated cytokines (IL‐23 and IL‐17) and granzyme B, in parallel with selectively increased IL‐10 expression in liver allografts. In vitro, galectin‐1 inhibited Flt3L‐differentiated DC‐mediated proliferation of allo‐CD4⁺ T cells and production of IFN‐γ and IL‐17. These data provide new evidence of the potential regulatory effects of galectin‐1 in alloimmune responses in a murine model of liver transplantation.     

Higher Risk of Kidney Graft Failure in the Presence of Anti‐Angiotensin II Type‐1 Receptor Antibodies

Reports have associated non‐HLA antibodies, specifically those against angiotensin II type‐1 receptor (AT1R), with antibody‐mediated kidney graft rejection. However, association of anti‐AT1R with graft failure had not been demonstrated. We tested anti‐AT1R and donor‐specific HLA antibodies (DSA) in pre‐ and posttransplant sera from 351 consecutive kidney recipients: 134 with biopsy‐proven rejection and/or lesions (abnormal biopsy group [ABG]) and 217 control group (CG) patients. The ABG's rate of anti‐AT1R was significantly higher than the CG's (18% vs. 6%, p < 0.001). Moreover, 79% of ABG patients with anti‐AT1R lost their grafts (vs. 0%, CG), anti‐AT1R levels in 58% of those failed grafts increasing posttransplant. With anti‐AT1R detectable before DSA, time to graft failure was 31 months—but 63 months with DSA detectable before anti‐AT1R. Patients with both anti‐AT1R and DSA had lower graft survival than those with DSA alone (log‐rank p = 0.007). Multivariate analysis showed that de novo anti‐AT1R was an independent predictor of graft failure in the ABG, alone (HR: 6.6), and in the entire population (HR: 5.4). In conclusion, this study found significant association of anti‐AT1R with graft failure. Further study is needed to establish causality between anti‐AT1R and graft failure and, thus, the importance of routine anti‐AT1R monitoring and therapeutic targeting.     

Improved transplantation outcome through delivery of DNA encoding secretion signal peptide‐linked glucagon‐like peptide‐1 into mouse islets

Glucagon‐like peptide‐1 (GLP‐1) stimulates cell proliferation and has anti‐apoptotic effects on pancreatic islet β cells. In our previous study, the transduction of mouse islets with a recombinant adenovirus containing GLP‐1 cDNA enhanced islet graft survival. In this study, we sought to deliver the GLP‐1 gene using a nonviral vector, which raises fewer safety issues in clinical application. We constructed a plasmid, pβ‐SP‐GLP‐1, in which a secretion signal peptide (SP) was inserted to increase GLP‐1 secretion, and transfected mouse islets using the nonviral carrier Effectene. Transfection of pβ‐SP‐GLP‐1 induced a significant increase in bioactive GLP‐1 levels in islet cultures. Islets transfected with pβ‐SP‐GLP‐1 were protected from H₂O₂‐induced cell damage in vitro. In addition, glucose‐stimulated insulin secretion was significantly increased in pβ‐SP‐GLP‐1‐transfected islets. Diabetic syngeneic mice transplanted under the kidney capsule with a marginal mass of pβ‐SP‐GLP‐1‐transfected islets rapidly became normoglycemic, with 88% of recipients being normoglycemic at 30 days post‐transplantation compared with 52% of mice that received pβ‐transfected islet grafts (P < 0.05). Islet grafts retrieved 7 days after transplantation revealed that the pβ‐SP‐GLP‐1‐transfected group had significantly more Ki67‐positive cells as compared with the pβ‐transfected group. In conclusion, delivery of a plasmid containing a secretion SP and GLP‐1 cDNA using a nonviral carrier leads to efficient secretion of GLP‐1 in mouse islet cells, enhances islet cell survival during the early post‐transplant period, and improves islet transplantation outcome.