High amounts of circulating interleukin (IL)-6 in the form of monomeric immune complexes during anti-IL-6 therapy. Towards a new methodology for measuring overall cytokine production in human in vivo.

A patient with plasma cell leukemia was treated with anti-interleukin (IL)-6 monoclonal antibodies (mAb) for 2 months. Using chromatography on protein A-Sepharose, anti-murine-IgG-Sepharose, anti-IL-6-mAb-Sepharose and gel filtration at pH 2.3, we have demonstrated that the anti-IL-6 mAb, by preventing the binding of IL-6 to its cell membrane receptor and its renal elimination, has induced huge amounts of IL-6 to circulate in the form of monomeric immune complexes. By using this observation, we have developed a mathematical modelling that allows the determination of the overall daily production of IL-6 in this patient, which was in the range of 15 micrograms per day. Overall in vivo production of cytokines has never been evaluated in animals or in humans before.     

Sexually dimorphic expression of protease nexin-1 and vanin-1 in the developing mouse gonad prior to overt differentiation suggests a role in mammalian sexual development

The mammalian sex-determining pathway is controlled by the presence or absence of SRY expression in the embryonic gonad. Expression of SRY in males is believed to initiate a pathway of gene expression resulting in testis development. In the absence of SRY, ovary development ensues. Several genes have now been placed in this pathway but our understanding of it is far from complete and several functional classes of protein appear to be absent. Sex-determining genes frequently exhibit sexually dimorphic patterns of expression in the developing gonad both before and after overt differentiation of the testis or ovary. In order to identify additional sex-determining or gonadal differentiation genes we have examined gene expression in the developing gonads of the mouse using cDNA microarrays constructed from a normalized urogenital ridge library. We screened for genes exhibiting sexually dimorphic patterns of expression in the gonad at 12.5 and 13.5 days post-coitum, after overt gonad differentiation, by comparing complex cDNA probes derived from male and female gonadal tissue at these stages on micro-arrays. Using in situ hybridization analysis we show here that two genes identified by this screen, protease nexin-1 (Pn-1) and vanin-1 (Vnn1), exhibit male-specific expression prior to overt gonadal differentiation and are detected in the somatic portion of the developing gonad, suggesting a possible direct link to the testis-determining pathway for both genes.     

Hyaluronan-related limited concentration by the immature kidney

The limited renal concentration performance by the immature kidney traditionally is thought to be attributed to blunted renal response to arginine vasopressin (AVP) and medullary hypotonicity. The diminished AVP-dependent osmotic water permeability of the collecting duct is the result of decreased AVP binding and adenylate cyclase activation, and low expression of aquaporin-2 (AQP2) mRNA and low levels of AQP2 protein. Moreover, the immature kidney fails to establish deep cortico-papillary osmotic gradient because of structural immaturity, limited solute transport and increased medullary blood flow. Based on indirect clinical and experimental evidences this article puts forward a hypothesis that during perinatal period the abundant hyaluronan (HA) content in the renomedullary interstitium has a primary role in antagonizing water reabsorption and limiting concentration performance. Hydration-related alterations in renal HA appears to be mediated by antidiuretic hormone.The concept of HA-mediated renal water transport may imply that interfering selectively with renal HA metabolism may provide a new therapeutic approach to promote diuresis or antidiuresis, respectively, according to the elevation or reduction in renomedullary HA.     

Needle necropsy in AIDS

The value of autopsy in understanding the natural course of any disease is beyond any argument. The reluctance of pathologists to perform autopsy in HIV infected cadavers is justified due to the risks involved to the prosector and the morgue attendants. A relative low risk needle necropsy protocol is proposed using fine needle aspiration cytology, tru-cut biopsies and microbiological examination. Diagnosis could be offered in all the forty-four needle necropsies performed. Disseminated tuberculosis in 18/44 (40.9%) cases, disseminated cryptococcosis in 12/44 (27.2%) cases, poly-microbial infections in 27.2% cases and non-Hodgkin's lymphoma in 9% cases were detected in the study. Infectious agents like Histoplasma capsulatum, Isospora belli, tachyzoites of Toxoplasma gondii, Candida sp and Cryptococcus sp could be demonstrated in the samples obtained in the study. Lack of material for study of gross pathology, inaccessibility of deep-seated lesions and risk of needle stick injury to the prosector though low are the limitations of this procedure.     

Mimotope and anti-idiotypic vaccines to induce an anti-IgE response

We have defined epitopes on human IgE by screening different phage display random peptide libraries with a monoclonal anti-IgE antibody termed BSW17. The selected mimotopes and epitopes within the Cepsilon3 and Cepsilon4 region of IgE induced antibodies that were nonanaphylactogenic and had biological activity similar to BSW17. The chemically synthesized and KLH-coupled IgE epitopes or mimotopes were used to induce an anti-IgE response in rhesus monkeys. The immunized rhesus monkeys were subsequently protected in a PCA test when sensitized with human IgE and triggered with the corresponding allergen. Furthermore, using the same monoclonal anti-IgE antibody, we also generated an anti-idiotypic antibody that showed sequence homology with the IgE epitope in the Cepsilon3 domain. This anti-idiotypic antibody as well as the mimotopes were then used in a mouse model to induce orally an anti-IgE immune response. For this purpose mice were fed by intragastric gavages with bacteriophages displaying the small IgE-homologous structures. Orally immunized mice produced serum anti-IgE antibodies that were inhibited by BSW17 suggesting that it may be possible to induce a systemic anti-IgE response orally.     

Study on the metastatic mechanisms of human giant-cell lung carcinoma comparison between the strains C and D

The biologic characteristics of the two human giant-cell lung carcinoma strains with high (strain D) and low metastatic potential (strain C) were studied, including karyotype of chromosome, intracellular free calcium ([Ca2+]i), morphologic changes of cell surface and the expression of nm23-H1, p53, ras, c-myc, c-erbB2, bcl-2 genes and PCNA. The correlation between different biologic features and the metastatic potential of the two strains was analyzed. We found: 1) Both strains had the karyotypic abnormality of -13, -14, -15, +20, +21 with seven same marker chromosomes. Only strain D had the karyotypic abnormality of +7, -17, -18, +X, 7p+; 2) [Ca2+]i of the strain C (984.7 +/- 573.8) and D (517.6 +/- 216.6) was significantly different (p < 0.05). The amplitude of intracellular calcium oscillations of strain C was lower than the one of strain D; 3) strain C had more villous-like protrusions on the cell surface, whereas strain D had more bubble-like protrusions; 4) The expression of nm23-H1 and p53 protein of strain C was all higher than that of strain D. The expression of PCNA of strain C was lower than strain D; 5) nm23-H1 mRNA levels of strain C was lower than that of strain D. We consider that the karyotype of chromosomes, intracellular free calcium, the structure of cell membrane and the expression of nm23-H1 gene, p53 gene, PCNA could be closely related to the metastatic potential of human giant-cell lung carcinoma. They could be used as the sign for judging whether the tumor will metastasize in clinical practice as well as in judging the prognoses of patients.     

A recombinant vaccinia virus expressing hepatitis B virus middle surface protein Restricted expression of HBV antigens in human diploid cells

Summary Several vaccinia virus recombinants inducing the synthesis of the middle surface (M) protein of hepatitis B virus (HBV) were constructed. One of them, denoted vl37, was examined in some detail. The virus replicated nearly to the same extent in various cell lines, viz. human embryo diploid fibroblast LEP and MRC-5 cells, rabbit embryo fibroblast REF cells, TK^– rat RAT-2 cells, and green monkey CV-1 cells. However, the production of M protein was found considerably lower in the human LEP and MRC-5 than in the other cells examined. In addition, the kinetics of M formation were different in these two cell systems, LEP cells lagging significantly behind CV-1 cells. The low-level production of M protein in LEP cells was not increased by repeated vl37 passages in LEP cells, nor by a passage in a laboratory worker accidentally infected with the vl37 virus, nor by shortening the leader sequence preceding the translation initiation codon. The greater part of the M antigen was found to be cell associated, more so in the cells of human than monkey origin. From the major HBV S antigen (HBsAg) isolated from the plasma of chronically infected subjects, the antigen released by cell destruction differed by binding to polymerized human albumin. This property was utilized in ELISA to detect anti-preS2 antibody. Rabbits inoculated intradermally with the vl37 virus developed antibodies reactive in this assay as well as with a synthetic peptide corresponding in the amino acids 14–34 of the NH₂terminus of the HBsAg preS2 region.     

Bacillus thuringiensis insecticidal delta-endotoxin: diversity of crystal proteins and its relatedness to the toxicity spectrum

Bacillus thuringiensis strains produce crystal delta-endotoxins which exhibit a diverse toxicity spectrum. In order to explore the basis of toxin specificity, a comparison of the activity of 13 strains belonging to seven serotypes was made against three insect species. The delta-endotoxin crystals were purified and their polypeptide composition analyzed by SDS-PAGE. Among the strains studied, the delta-endotoxins consist of a variety of crystal proteins in the 60-144 KDa size range. On the basis of molecular mass, endotoxins maybe grouped into two classes; one contained both high (125-144 KDa, P1) and medium sized (60-66 KDa, P2) proteins and a second class consisting of only the high Mr polypeptides. Immunoblotting with B. aizawai P1 antiserum revealed antigenic cross-reaction with one or more of the polypeptides in 125-144 KDa range in all the strains studied. When the crystal proteins from different strains were immunoblotted with kurstaki P2 antiserum, none of the P1 protein crossreacted suggesting that the P1 and P2 proteins are not structurally related. However, the B. kurstaki P2 antiserum crossreacted with 66 KDa proteins in some other strains which underlines a structural homology in this class of the toxic polypeptides. Toxicity studies revealed that the high Mr P1 proteins of all the strains in this study were active against lepidopteran (Pieris brassicae and Diacrisia obliqua) larvae. B. thuringiensis aizawai strains exhibit a dual toxicity associated with the high Mr (130-135 KDa; P1) proteins. The P2 crystal proteins (60-66 KDa) also showed dual toxicity against the lepidopteran and dipteran larvae but were found to be structurally and immunologically distinct.     

Late asthmatic response to allergen challenge (LAR), its clinical feature and pharmacological modulation

Conclusion The LAR can regularly be associated with late skin response, increased serum IgG, blood eosinophilia and late bronchial complaints. The LAR is prevented highly by BDA (BSA) and DSCG. The exact mechanism underlying the LAR is not yet fully known [2–4]. However, with respect to this study, the involvement of the type III hypersensitivity in the clinical LAR could be presumed to be one of the possible mechanisms [2, 3].     

Clinical effect of hypoallergenic rice (HRS-1) in atopic dermatitis. HRS-1 Research Group

The usefulness of hypoallergenic rice (HRS-1) was clinically evaluated in 43 patients with severe atopic dermatitis (AD), who were suspected of having rice allergy, in collaboration with 13 hospitals. The patients were fed with HRS-1 instead of eliminating both regular rice and wheat from their daily diet. AD area and severity index (ADASI) was calculated as an indicator of the degree of cutaneous symptoms. Significant decrease of ADASI were observed in the 2nd and 4th week and at the end of the replacement therapy (5.6 weeks on average). On final evaluation, 74% of the patients tested showed "moderate" to "remarkable" improvement, and in 53% of the patients HRS-1 resulted in a "moderate" to "remarkable" reduction in the dosage and the grade of potency of the steroid ointment concomitantly used for the treatment. Finally, HRS-1 was evaluated as "useful" to "very useful" as the elimination diet in 70% of the patients.