Hyperthermic treatment of chromomycosis with disposable chemical pocket warmers

A case of chromomycosis in which hyperthermia proved effective is reported. The patient was a 56-year-old male bean curd maker who, without any previous history of minor trauma, developed on the extensor side of the left upper arm an eczematous lesion that underwent gradual radial expansion. The lesion showed a well-defined, 7×10 cm infiltrated erythematous plaque with the central area healed and, at the upper and lower borders, adherent scales and crusts on the surface. Histological examination revealed granulomatous changes in the dermis, as well as sclerotic cells within giant cells and microabscesses. On culturing,Fonsecaea pedrosoi was isolated. The patient was treated with disposable chemical pocket warmers, which were secured over the lesion with a rather tight elastic bandage, so that they kept the affected area warm for 24 hours a day. After a month of such hyperthermic treatment, the erythema and infiltration had decreased considerably, and microscopic examination and culture of the crusts both yielded negative results. Examination of biopsy specimens of the lesion after the third month showed that it had cicatrized. The treatment was stopped after 4 months, and no relapse occurred. We also summarize the published results of local hyperthermic treatment of chromomycosis in Japan.     

Identification of HLA-B44 subtypes associated with extended MHC haplotypes

The HLA class I antigen B44 is found in each of two different extended major histocompatibility haplotypes (allele combinations of HLA-B, HLA-DR, and complement genes BF, C2, C4A, and C4B in linkage disequilibrium). Using isoelectric focusing, two variants of HLA-B44 were identified. The basic variant was found in all cell lines with the extended haplotype HLA-B44, DR7, FC31, and the acidic variant in all cell lines with the extended haplotype HLA-B44, DR4, SC30. The occurrence of each antigen variant with a unique extended haplotype explains previous observations concerning the nonrandom association of B44 variants with DR antigens.     

Tissue-specific regulation of GTP-binding protein and muscarinic acetylcholine receptor levels during cardiac development

A quantitative immunoblot assay was developed by using affinity-purified monospecific antibodies to quantitate levels of guanine nucleotide binding regulatory protein (G-protein) subunits in atria and ventricles during embryonic chicken cardiac development. The muscarinic acetylcholine receptor (mAChR) number was measured with [3H]quinuclidinyl benzilate. On day 10 of embryonic development (day 10E) there was no difference between the atrial and ventricular membrane concentrations of beta-subunit, G0 alpha subunit, or mAChR. The level of Gi alpha was found to be 44% greater in atria than in ventricles on day 10E. The atrial membrane concentration of beta-subunit increased 80% between day 13E and 15E, G0 alpha increased 46% between day 10E and 15E, mAChR increased 61% between day 10E and 12E, and Gi alpha decreased 34% between day 10E and 13E. The atrial levels of beta-subunit, G0 alpha, Gi alpha, and mAChR did not change further through day 20E. The ventricular membrane concentration of these proteins did not change between day 10E and 20E, except for that of G0 alpha, which increased 47% between day 15E and 20E. The atrial specific increase in beta-subunit correlated with a loss of GTP inhibition of basal adenylate cyclase activity. The difference in Gi alpha levels between atria and ventricles on day 10E correlated with a difference in carbachol sensitivity of atrial and ventricular basal adenylate cyclase activity. Thus, the levels of several components of the cholinergic neuroeffector pathway are regulated in a tissue-specific manner at a time that coincides with the onset of functional parasympathetic innervation of the embryonic chicken heart.(ABSTRACT TRUNCATED AT 250 WORDS)     

Chemically induced hypothyroidism produces elevated amounts of the alpha subunit of the inhibitory guanine nucleotide binding protein (Gi) and the beta subunit common to all G-proteins.

Adipocytes of hypothyroid rats display an increased responsiveness to agents which function by inhibiting the production of cyclic AMP. Anti-peptide antisera which selectively recognise the alpha subunit of the inhibitory guanine nucleotide binding protein (Gi) detected a 40 kDa polypeptide in adipocyte plasma membranes of both euthyroid and hypothyroid rats. Amounts of the alpha subunit of Gi were elevated some 2-fold in the hypothyroid preparations in comparison with the euthyroid controls, when equal amounts of membrane protein of the two treatments were examined. As cells from the hypothyroid animals contained 2.7 times as much membrane protein as those from the control animals, the amounts of alpha subunit of Gi are elevated some 5.6-fold per cell in adipocytes of the hypothyroid animals compared with the euthyroid controls. Amounts of the 36 kDa beta subunit of G-proteins were also elevated in plasma membranes of adipocytes of hypothyroid animals, in this case by some 50% when compared on a protein basis. These results provide direct evidence for alterations in the amounts of the subunits of Gi caused by the hypothyroid state.     

Persistent activation of the alpha subunit of Gs promotes its removal from the plasma membrane.

As assessed both by cholera-toxin-catalysed ADP-ribosylation and by immunoblotting with an anti-peptide antiserum raised against the C-terminal decapeptide of forms of Gs alpha (the alpha subunit of the stimulatory guanine nucleotide-binding protein), rat glioma C6 BU1 cells express two forms of Gs alpha: a major 44 kDa form and a much less prevalent 42 kDa form. We examined the effects of guanine nucleotides on the interaction of the 44 kDa form with the plasma membrane. Incubation of membranes of C6 BU1 cells with poorly hydrolysed analogues of GTP, but not with analogues of either ATP or GDP, caused the release of this Gs alpha from the membrane fraction. Release of Gs alpha was observed within 5 min, and continued throughout the incubation period. After treatment with guanosine 5'-[beta gamma-imido]triphosphate for 60 min, some 75% of this polypeptide had been released from its site of membrane attachment. These experiments demonstrate that Gs alpha need not remain associated invariantly with the plasma membrane.     

Cholera toxin treatment produces down-regulation of the alpha-subunit of the stimulatory guanine-nucleotide-binding protein (Gs).

The effect of activation of the alpha-subunit(s) of the stimulatory guanine-nucleotide-binding protein, Gs, on levels of this polypeptide(s) associated with the plasma membrane of L6 skeletal myoblasts was ascertained. Incubation of these cells with cholera toxin led to a time- and concentration-dependent 'down-regulation' of both 44 and 42 kDa forms of Gs alpha as assessed by immunoblotting with an anti-peptide antiserum (CS1) able to identify the extreme C-terminus of Gs. The effect of cholera toxin was specific for Gs; levels of Gi alpha in membranes of cholera toxin-treated cells were not different from untreated cells. Down-regulation of Gs was absolutely dependent upon prior ADP-ribosylation, and hence activation of Gs and was not mimicked by other agents which elevate intracellular levels of cyclic AMP. Pretreatment with pertussis toxin, which catalyses ADP-ribosylation of Gi but not of Gs, did not down-regulate either Gi or Gs, demonstrating that covalent modification by ADP-ribosylation is alone not a signal for removal of G-proteins from the plasma membrane.     

Antibodies which recognize the C-terminus of the inhibitory guanine-nucleotide-binding protein (Gi) demonstrate that opioid peptides and foetal-calf serum stimulate the high-affinity GTPase activity of two separate pertussis-toxin substrates.

We investigated the mechanisms of receptor-mediated stimulation of high-affinity GTPase activity in response to opioid peptides and to foetal-calf serum in membranes of the neuroblastoma X glioma hybrid cell line NG108-15. Increases in GTPase activity in response to both of these ligands was abolished by prior exposure of the cells to pertussis toxin. Pertussis toxin in the presence of [32P]NAD+ catalysed incorporation of radioactivity into a broad band of approx. 40 kDa in membranes prepared from untreated, but not from pertussis-toxin-pretreated, cells. Additivity studies indicated that the responses to opioid peptides and to foetal-calf serum were mediated by separate guanine-nucleotide-binding proteins (G-proteins). Whereas opioid peptides produced an inhibition of adenylate cyclase in membranes of untreated cells, foetal-calf serum did not. Affinity-purified antibodies which recognize the C-terminus of the inhibitory G-protein identified a 40 kDa polypeptide in membranes of NG108-15 cells. These antibodies attenuated opioid-stimulated high-affinity GTPase activity, but did not markedly affect the response to foetal-calf serum. We conclude that receptors for the opioid peptides function via the inhibitory G-protein (Gi), whereas foetal-calf serum activates a second pertussis-toxin-sensitive G-protein, which has a C-terminal sequence significantly different from that of Gi.     

Differential regulation of amounts of the guanine-nucleotide-binding proteins Gi and Go in neuroblastoma x glioma hybrid cells in response to dibutyryl cyclic AMP.

Incubation of the neuroblastoma x glioma hybrid cell line NG108-15 in tissue culture with dibutyryl cyclic AMP (1 mM) for up to 8 days produced a morphological differentiation of the cells, during which they extended neurite-like processes. Pertussis-toxin-catalysed ADP-ribosylation indicated that amounts of guanine-nucleotide-binding proteins (G-proteins), which are substrates for this toxin, were approximately doubled in membranes from the 'differentiated' cells in comparison with the control cells. Immunoblotting of membranes derived from either untreated or dibutyryl cyclic AMP-treated cells with anti-peptide antisera specific for the alpha subunits of the pertussis-toxin-sensitive G-proteins Gi and Go demonstrated that amounts of these G-proteins were reciprocally modulated during the differentiation process. In comparison with the untreated cells, the amount of Gi in the 'differentiated' cells was decreased, whereas the amount of Go was substantially increased. Stimulation of high-affinity GTPase activity in response to opioid peptides, which in this cell line interact with an opioid receptor of the delta subclass, was much decreased, and inhibition of adenylate cyclase activity was almost entirely attenuated in the 'differentiated'-cell membranes in comparison with membranes of untreated cells. Opioid receptor number was also decreased in membranes of the dibutyryl cyclic AMP-treated cells in comparison with the control cells. These data demonstrate that relatively small changes in the observed pattern of pertussis-toxin-catalysed ADP-ribosylation of membranes can mask more dramatic alterations in amounts of the individual pertussis-toxin-sensitive G-proteins, and further demonstrate the importance of methodologies able to discriminate between the different gene products.     

Pulmonary paracoccidioidomycosis in immunized mice

Paracoccidioidomycosis was induced in immunized (IM) and non-immunized (NI) mice. The histopathology, the number of fungi in the lungs, the cellular (footpad test — FPT and macrophage inhibition factor assay — MIF) and humoral (immunodiffusion test) immune response were investigated serially postinfection. In the IM mice, at days 1 and 3, there was intense and predominant macrophagic-lymphocytic alveolitis with loose granulomatous reaction; at day 30, inflammation was mild. In the NI group, up to day 3, the lesions were focal; later there was formation of extensive epithelioid granuloma. The number of fungi in IM mice were always smaller than those of NI group. Immunization alone induced positive FPT and MIF indices with low titer of antibody. After infection, there was a significant decrease of the FPT indices in the IM group, which we interpreted as desensitization due to trapping of sensitized lymphocytes in the lungs. In conclusion, (1) The lesional pattern of pulmonary paracoccidioidomycosis in IM mice was similar to that of a hypersensitivity pneumonitis. This reaction was probably effective in reducing the extension of the infection and decrease the number of fungi. (2) In this model, pulmonary resistance against P. brasiliensis seems to be related to local and systemic delayed-type hypersensitivity reaction.