Bullous pemphigoid (BP) is characterized by autoantibodies against 230- and 180-kDa hemidesmosomal antigens located in the most superficial layers of the basement membrane zone (BMZ). Histologically, there is a predominance of eosinophils in the infiltrate. In a psoriatic patient, we identified an unusual autoimmune subepidermal bullous eruption which clinically resembled BP, but which was characterized by IgG autoantibodies against a novel 200-kDa lower lamina lucida component. Histologically there was a predominance of neutrophils in the infiltrate. Direct immunofluorescence showed linear immunoglobulin (Ig)G and C3 deposition at the BMZ. The patient's IgG autoantibodies bound exclusively to the dermal side of salt-split normal human skin. Indirect immunogold electron microscopy showed a marked deposition of IgG at the lower lamina lucida and minimal deposition at the hemidesmosomes. Immunoblot analysis identified a unique 200-kDa autoantigen in dermal extracts and a faint band of the 230-kDa BP antigen in epidermal extracts. The patient responded dramatically well to cyclosporin A. Although the patient's serum also reacted slightly with the 230-kDa BP antigen, there were significant findings different from the usual immunopathological changes of BP. These included finding a novel 200-kDa lower lamina lucida target antigen, the binding of IgG autoantibodies exclusively to the dermal side of the split skin and a predominance of neutrophils in blister infiltrate. The IgG autoantibodies against the 200-kDa lamina lucida target antigen seemed to play a major role in the pathogenesis of this unique autoimmune subepidermal dermatosis.

Exposure of human keratinocyte HaCaT cells to ultraviolet B-irradiation induced apoptotic morphologic changes. In this study, we found that the ultraviolet B irradiation (0.25 J per cm2) induced phosphorylation of p38 mitogen-activated protein kinase and c-jun N-terminal protein kinase, and also significant activation of caspase-3 (CPP32-like protease) and a small increase of caspase-1 (ICE-like protease) activity in the early stages of ultraviolet B-induced apoptosis. Pretreatments of the cells with a p38 mitogen-activated protein kinase inhibitor, SB203580, and a caspase-3 inhibitor, Ac-Asp-Met-Gln-Asp-1-aldehyde, suppressed the ultraviolet B irradiation-induced apoptosis by approximately 60% as estimated by nuclear staining and DNA laddering. Pretreatment with caspase-1 inhibitor, Ac-Tyr-Val-Lys-Asp-aldehyde was without effect. Ultraviolet B-induced caspase-3 activation resulted in cleavage of poly(ADP) ribose polymerase, which was abolished by the caspase-3 inhibitor. SB203580 pretreatment prevented activation of caspase-3 and caspase-1, and also suppressed the cleavage of poly(ADP) ribose polymerase. Neither ceramide generation nor sphingomyelinase activation (neutral and acid) was observed in the ultraviolet B-irradiated HaCaT cells. Also various antioxidants did not affect the caspase activation induced by ultraviolet B irradiation. These results indicated that activation of p38 mitogen-activated protein kinase upstream of caspases may play an important part in the apoptotic process of keratinocytes exposed to ultraviolet B irradiation.

We performed an electrophysiological study to investigate ion transport of pendrin and thereby understand the pathogenesis of Pendred syndrome. Using pendrin-transfected COS-7 cells, we could show that pendrin transports both iodide and chloride measured as voltage-dependent inward and outward membrane currents. Chloride in the culture medium, [Cl-]o, was efficiently exchanged with cytoplasmic iodide, [I-]i, under physiological concentrations, indicating that pendrin is important for chloride uptake and iodide efflux. Although exchange of iodide in the medium, [I-]o, with cytoplasmic chloride, [Cl-]i, was observed, a significantly high concentration of iodide (10 mm) was required. In addition, either iodide or chloride was required on both sides of the cell membrane for the anion exchange activity of pendrin, indicating that iodide and chloride activate the exchange activity of pendrin while they are transported. The present study further supports that pendrin is responsible for the iodide efflux in thyroid cells where intracellular iodide concentration is high and that the general function of pendrin in other tissues is to transport chloride through exchange with other anions.

RNA interference by short interfering RNAs (siRNAs) holds promise as a therapeutic strategy, but use of siRNAs in vivo remains limited. Here, we developed a system to target delivery of siRNAs to glomeruli via poly(ethylene glycol)-poly(L-lysine)-based vehicles. The siRNA/nanocarrier complex was approximately 10 to 20 nm in diameter, a size that would allow it to move across the fenestrated endothelium to access to the mesangium. After intraperitoneal injection of fluorescence-labeled siRNA/nanocarrier complexes, we detected siRNAs in the blood circulation for a prolonged time. Repeated intraperitoneal administration of a mitogen-activated protein kinase 1 (MAPK1) siRNA/nanocarrier complex suppressed glomerular MAPK1 mRNA and protein expression in a mouse model of glomerulonephritis; this improved kidney function, reduced proteinuria, and ameliorated glomerular sclerosis. Furthermore, this therapy reduced the expression of the profibrotic markers TGF-␤1, plasminogen activator inhibitor-1, and fibronectin. In conclusion, we successfully silenced intraglomerular genes with siRNA using nanocarriers. This technique could aid the investigation of molecular mechanisms of renal disease and has potential as a molecular therapy of glomerular diseases.

PA28-gamma is the activator of 20S proteasome, the ATP-dependent proteolytic system that plays an important role in cell cycle progression in various cell types. In this paper, we show the abnormally high expression of PA28-gamma in various thyroid neoplasms. Thyroid samples were obtained from patients with normal thyroid (4 cases) and with the following diseases: papillary adenocarcinoma (13 cases), multinodular goiter (4 cases), and anaplastic carcinoma (1 case). PA28-gamma expression was estimated by immunohistochemical staining and Western blotting. In all of the papillary adenocarcinoma samples, PA28-gamma was abnormally overexpressed, especially in cancer cells existing at the peripheral region of the cancer mass or in cancer cells invading the capsular region surrounding the cancer mass. In cancer cells of these areas, PA28-gamma was predominantly distributed in nucleus rather than in the cytoplasm of cancer cells. On the other hand, no obvious PA28-gamma expression was observed in the adjacent normal thyroid follicular cells. In multinodular goiter, the expression of PA28-gamma was relatively low compared with papillary adenocarcinoma. In anaplastic carcinoma, PA28-gamma was expressed at the highest level, especially in poorly differentiated regions such as squamous metaplasia of anaplastic cancer tissue. Therefore, the PA28-gamma expression seems to be restricted to thyroid cancer cells, especially in the region where the growth rate of cancer cells is accelerated. This result is further confirmed by the fact that C2, alpha-subunit of 20S proteasome, and proliferating cell nuclear antigen are similarly overexpressed in this region. Thus, PA28-gamma might be involved in the regulatory system for the cell cycle. Moreover, the growth of thyroid cancer cell lines was affected by the proteasome inhibitor, clasto-lactacystin beta-lactone. These results demonstrate that PA28-gamma is overexpressed in thyroid cancer, especially in its growth-accelerated cells.