Efficacy of Cryofiltration for Treatment of Mixed Cryoglobulinemia: A Report of Four Cases.

Cryoglobulinemia can induce systemic vasculitis affecting various organs such as skin, peripheral nerves, and kidney. The disease can induce chronic organ failure and even be life-threatening. Cryofiltration has been applied for the treatment of cryoglobulinemic vasculitis. We have experienced four cases with mixed cryoglobulinemia showing severe and progressive clinical manifestations, including skin purpura, nephrotic syndrome, acute kidney injury, and peripheral neuropathy. Cryofiltration in conjunction with conventional pharmacological therapies appeared to be safe and effective. After the treatments, plasma cryoglobulins were markedly reduced and the disease was well controlled. Although its efficacy has not yet been well established, this report can be another evidence showing efficacy of cryofiltration for treatment of mixed cryoglobulinemia.

[Case of anti-glomerular basement membrane antibody nephritis in a patient who was able to withdraw from dialysis and gave birth twice but underwent living renal transplantation due to progression to end-stage renal failure 15 years after onset].

A 34-year-old woman with suspected rapidly progressive glomerulonephritis had been admitted to our hospital in March 1993 at the age of 19 years. Renal biopsy revealed cellular crescent formation in 24 of 26 glomeruli. Serum examination was positive for anti-glomerular basement membrane (GBM) antibody, while pulmonary hemorrhage was absent. Based on these findings, she was diagnosed with anti-GBM antibody nephritis, and treated with corticosteroid pulse therapy and double filtration plasmapheresis (DFPP) in addition to hemodialysis (HD). HD was withdrawn within 2 months. Wishing to have a baby, she had delivery in 1997 and 2000. Subsequently, her renal function gradually decreased, and she underwent an ABO-incompatible living-donor kidney transplant, with her husband as the donor, in March 2008. She has been making good progress after transplantation. Anti-GBM antibody nephritis has a poor prognosis, but renal function was maintained for 15 years in this patient, who responded well to the initial treatment. The underlying disease rarely recurs if transplantation is performed after the patient has become negative for anti-GBM antibody, anti-GBM antibody nephritis therefore seems to be a good indication for treating patients with renal transplantation.

Induction of pulmonary thromboembolism by neutrophil elastase in collagen-induced arthritis mice and effect of recombinant human soluble thrombomodulin.

We previously reported that during total knee arthroplasty in rheumatoid arthritis (RA) patients, the use of tourniquet might promote local release of neutrophil elastase (NE) from neutrophils, which may contribute to the development of pulmonary thromboembolism (PTE) and tissue injury. The aim of this study was to develop PTE by the use of NE in a mouse model of collagen-induced arthritis (CIA) and investigate the relationship between thrombus and endothelial cells as well as the effect of recombinant human soluble thrombomodulin (rhs-TM) in reducing the risk of PTE. Male DBA/1J mice were injected intracutaneously at several sites with an emulsion containing bovine collagen and later a booster shot to produce CIA mice. Subsequently, NE was injected intravenously 2 times a day for 3 days and after a further 4 days, mice were sacrificed. A group of mice received rhs-TM injections prior to NE injections. We divided the mice into four groups of normal, CIA control, CIA + NE, and CIA + rhs-TM + NE mice and evaluated thrombus formation status. All CIA + NE mice developed PTE. In contrast, no thrombosis was found in normal control, CIA control and CIA + rhs-TM + NE mice. Plasma thrombin level, fibrinogen expression and neutrophil count were increased in CIA + NE mice. Double staining for anticoagulant TM and procoagulant von Willebrand factor (vWF) in pulmonary endothelial cells in normal mice showed a TM-dominant expression while in both CIA control and CIA + NE mice a vWF-dominant expression compatible with coagulant status was observed. Injection of rhs-TM into CIA + NE mice resulted in a phenotypic conversion of endothelial cells from vWF-dominant to TM-dominant expression and a reduction in fibrinogen deposition. These findings demonstrate that by repeated use of NE in CIA mice, it is feasible to produce PTE and to study its pathogenesis and that rhs-TM reduces the risk of PTE. We suggest that in surgical operations of upper and lower extremities in RA patients, the use of a tourniquet should be avoided as it may trigger NE release.

Hepatocyte growth factor prevents peritoneal fibrosis in an animal model of encapsulating peritoneal sclerosis.

BACKGROUND: Ultrafiltration failure associated with peritoneal fibrosis can lead patients to discontinue continuous ambulatory peritoneal dialysis (CAPD). It has been reported that the reciprocal imbalance between transforming growth factor-beta1 (TGF-beta1) and hepatocyte growth factor (HGF) is closely involved in the progression of tissue fibrosis. We previously showed that exogenous HGF restores the growth of human peritoneal mesothelial cells suppressed by a high concentration of D-glucose or TGF-beta1. In this study, we examined whether constitutive exposure to HGF prevents peritoneal fibrosis in an animal model of encapsulating peritoneal sclerosis (EPS). METHODS: To establish the model, a daily intraperitoneal injection of 0.1% chlorhexidine gluconate was given to male Wister rats for 35 days. Rat peritoneal mesothelial cells (RPMCs) transfected with full-length human HGF cDNA in an expression vector (pUCSRalpha/HGF) were injected into the peritoneal cavity of the rats. Thereafter, pathological changes to the peritoneal membrane were observed, and the effect on peritoneal ultrafiltration volume was examined. RESULTS: In the model, microscopic examination revealed a progressive thickening of the submesothelial layer, and an increase in the number of capillary vessels. Peritoneal ultrafiltration volume was decreased. Interestingly, the pathological changes to the peritoneal membrane were reversed by the intraperitoneal injection of pUCSRalpha/HGF-transfected RPMCs. Furthermore, peritoneal ultrafiltration volume was increased. CONCLUSIONS: The constitutive production of HGF by UCSRalpha/HGF-transfected RPMCs can improve peritoneal fibrosis resulting in an increase in peritoneal ultrafiltration volume. This approach may have clinical application.

Critical roles of capillary endothelial cells for alveolar remodeling in nonspecific and usual interstitial pneumonias.

To characterize the relationship between angiogenesis factors and alveolar remodeling in interstitial lung diseases, we examined alveolar capillary endothelial cells in the normal lung (n=5) and in lungs with nonspecific interstitial pneumonia (NSIP) (n=4) or usual interstitial pneumonia (UIP) (n=6) using immunofluorescence staining for thrombmodulin and von Willebrand factor (vWF). With three-dimensional images of alveolar capillaries, the diameter of capillary tubes and their branching frequency per unit length were determined to define rearrangement of the capillary meshwork. Alveolar capillary endothelial cells in normal lungs expressed surface thrombomodulin, and those in lungs with cellular NSIP often showed coexpression of surface thrombmodulin and cytoplasmic vWF. In the alveolar septa of fibrotic NSIP and UIP, capillary endothelial cells demonstrated vWF in only the cytoplasm. Capillary branching frequencies in NSIP and UIP were decreased to 45% and 22%, respectively, of the normal level (p<0.002). Compared with normal lungs, in NSIP and UIP lungs alveolar capillaries containing TUNEL-positive endothelial cells (p<0.05) showed increases of 3.6-fold and 4.3-fold, respectively, indicating a close correlation between endothelial cell apoptosis and remodeling of alveolar capillary frameworks. The analysis of mRNA expression of vascular endothelial growth factors (VEGF) and their receptors (VEGFR1 and VEGFR2) showed a significant decrease in each VEGF isoform and in VEGFR2 mRNA in representative alveolar wall tissues microdissected from the normal, NSIP, and UIP lungs. These results suggest that decreased expression of VEGF mRNA is associated with a reduction in the number of capillary tubes via endothelial cell apoptosis that possibly results in alveolar remodeling in NSIP and UIP. However, whether VEGF is related to fibroblastic activation in the interstitial matrix remains unclear.

Possible molecular target therapy with rapamycin in MDS.

The authors previously reported the mRNA expression of Glutathione S-transferases theta (GSTT)-1, wild type (623 bp) and mutant (500 bp) in MDS patients. The deletion of 123 bp creates a sequence that is homologues to mammalian target of rapamycin (mTOR). To analyse the function of mutant GSTT-1 gene, stable transformants for the mutant and wild-type GSTT-1 gene, respectively, were established. In this study, the expression of wild and mutant type GSTT-1 gene of those stable transformants and bone marrow cells from MDS patients by RT-PCR was observed in the presence or absence of rapamycin. In result, exposure of rapamycin led to the disappearance of just the mutant gene band. This phenomenon possibly indicates that rapamycin only attacked the mutant GSTT-1 expressing clone.

Possible effects of hepatocyte growth factor for the prevention of peritoneal fibrosis.

OBJECTIVE: Some patients who had carried out long-term continuous ambulatory peritoneal dialysis discontinued the treatment because of progressive peritoneal fibrosis. It has been previously reported that transforming growth factor-beta1 (TGF-beta1) is one of the factors that induces peritoneal fibrosis. Also, hepatocyte growth factor (HGF) plays a role in the prevention of fibrosis and in inhibiting TGF-beta1 production. In this study, we examined the effects of HGF on peritoneal fibrosis by TGF-beta1 induced by high concentrations of D-glucose. DESIGN: We transfected a full-length human HGF cDNA in an expression vector into human peritoneal mesothelial cells (HPMCs) using the calcium phosphate method. Transfected HPMCs were cultured with high concentrations of D-glucose solution and co-cultured with fibroblasts using a transwell system. Cell proliferation was determined using the Tetra Color One method. TGF-beta1 and HGF protein were measured by enzyme-linked immunosorbent assay. RESULTS: In addition to recombinant HGF, the growth inhibition of HPMCs by high concentration D-glucose or TGF-beta1 was significant. By transfecting HGF cDNA into HPMCs, growth inhibition by high concentration D-glucose was completely restored. Furthermore, the production of TGF-beta1 was also significantly decreased. CONCLUSION: These results suggested that exogenous HGF could possibly prevent peritoneal fibrosis.

Demonstration of microvessel networks and endothelial cell phenotypes in the normal murine lung.

The vascular endothelial cells (ECs) express various antigens related to coagulation factors, including factor VIII-related antigen or von Willebrand factor (vWF) in the cytoplasm and thrombomodulin (TM; a thrombin receptor)along the plasma membrane. CD34 (a hematogenic stem cell marker) is also expressed along the surface membrane of the ECs. Using these EC markers and fluorescein-isothiocyanate-labeled dextran (FITC-dextran)(Sigma Co., St. Louis, MO), we attempted to demonstrate the complex network of microvessels and their EC phenotypes in tracheo-bronchial trees and lung parenchyma of the normal adult ICR male mice. Under anesthesia, saline with heparin was infused slowly through left ventricle to drain off the blood. Following brief fixation with 4% buffered paraformaldehyde solution (PFA) through the same route, one group of animals received, 1) FITC-dextran injection via left ventricle, and the large airways and lungs were further fixed in PFA, or 2) The airways and lungs of the other group were rapidly frozen, and the thin sections were stained with two antibodies of vWF and Alexa Fluor 594-labeled CD34. The vWF antibody was later labeled by FITC. The microvessels of airways and lungs were observed by a laser scanning confocal microscope (TC-SP, Leica, Heidelberg, Germany). The phenotypic characteristics of microvessel ECs appeared mostly identical with those described previously in the human lung, although CD34 was applied instead of TM in the present study. The topographical heterogeneity of immunohistochemical properties of ECs would suggest functional differences at different sites of the lung, that would provide a novel insight for understanding the pathogenesis of human lung diseases.