Hypersensitivity to antigen derived from brain tissue in patients with Huntington's disease and in individuals at risk.

The hypothesis that an immune response is directed to components of brain tissue in Huntington's disease (HD) was examined. 27 patients with HD, 21 children of affected parents who showed no signs of disease, 6 spouses of HD patients, and 40 patients with other neurologic diseases were investigated. Leukocyte migration inhibition to antigens prepared from brain tissue was demonstrated in 26 HD patients and in 9 children. None of the six spouses reacted. 13 patients with other neurologic conditions (including 60% of patients after cerebrovascular accidents) reacted to rat brain extract. Fractionation of brain tissue indicated that the material responsible for reactions in these patients differed from that in HD. The material that elicited hypersensitivity in HD was not species specific, since it was found in rat, calf, human, and rabbit brain. It was not present in rat kidney, liver, or heart. Preliminary characterization showed that the material was protein in nature, contained sugar and lipid, and was heat labile. The finding of hypersensitivity in about half the unaffected children of HD parents suggests that it may precede the clinical onset of disease and serve as a marker for the gene.

Gland to gland heterogeneity in histologically normal mucosa of colon cancer patients demonstrated by monoclonal antibodies to tissue-specific antigens.

Two of three monoclonal antibodies to tissue-specific antigens of isolated colonic glands revealed gland to gland heterogeneity of antigen expression in sections of the histologically normal colonic mucosa from patients with colorectal adenocarcinoma or various nonmalignant conditions. A colon-specific goblet cell antigen (designated 3NM) was absent from rare, solitary glands randomly distributed among the otherwise strongly stained glands of distal colon. These 3NM-negative glands stained normally for the other two antigens studied and appeared morphologically and histochemically normal. They occurred in 11 of 13 cancer patients and in each of the five patients with benign conditions with median incidences of 2.2 and 0.4 per 1000 glands, respectively. Gland heterogeneity was also demonstrated in both patient groups for a cell membrane antigen (designated 6NM). In cecum and ascending colon, glands staining strongly and weakly for 6NM were found intermixed. The weakly stained glands tended to predominate in cecum and first part of ascending colon, but they were completely replaced by strongly stained glands in more distal colon. The heterogeneity shown by both 3NM and 6NM appeared due to phenotypically distinct cell clones. Our observations indicate that histologically normal colonic mucosa contains antigenically diverse gland populations.

Antigen expression in normal and neoplastic colonic mucosa: three tissue-specific antigens using monoclonal antibodies to isolated colonic glands.

Monoclonal antibodies to normal colonic glands were used to localize three tissue-specific antigens in sections of 40 hyperplastic polyps, 29 adenomas, 43 colorectal adenocarcinomas, and blocks or rolls of histologically normal large bowel mucosa. Two antibodies identified antigens (designated 3NM and 17NM) associated with the mucin vacuole of goblet cells and the third antibody identified a cell membrane antigen (designated 6NM). Antigen 3NM was absent from virtually all carcinoma cells and from all, or most, cells of hyperplastic polyps. Antigen 17NM was absent from 21 carcinomas, with some antigen-positive cells present in the others. Antigen 6NM content was unchanged compared to normal mucosa in 12 carcinomas, decreased somewhat in another 20, and either absent or markedly depleted in 11. Antigen 6NM increased in 22 hyperplastic polyps. Three cancer patients synthesized 6NM in their normal mucosa but the cellular distribution was abnormal. Antigens decreased in most adenomas, with heterogeneity of expression seen in glands and cells of some adenomas. Low concentrations of one or more antigens in normal mucosa were significantly more frequent (P less than 0.05) in patients with metastases than in those with stage A or B tumors.

Molecular cloning and characterization of a cDNA encoding the human leucocyte vacuolar protein sorting (h1Vps45).

We have isolated a novel cDNA clone from human leucocyte cDNA library, encoding a Sec1p-like vacuolar protein sorting (h1Vps45) which is believed to be implicated in vesicular transportation. Although the deduced amino acid (AA) sequence of this cDNA has revealed 97% identity to other known mammalian vacuolar protein sorting, there is an extensive variation in nucleotide sequence in comparison to that of three previously reported human (hVps45), rat (rVps45) and mouse (mVps45) vacuolar protein sorting (Vps45) cDNAs [1-3]. At the nucleotide sequence level h1Vps45 demonstrated 90% homology to the hVps45 and rVps45 and 89% identity to mVps45 with no significant homology in their noncoding regions. The 2.4 Kb mRNA corresponding to the h1Vps45 clone is widely distributed in a variety of human tissues expressing highest levels in peripheral blood mononuclear cells (PBMC), neutrophils, heart, spleen, and testis. The chromosomal mapping studies have demonstrated that the h1Vps45 is localized to long arm of human chromosome 1 at q21-q22. Our data indicates that we have isolated, characterized and mapped a novel cDNA encoding h1Vps45, which may play an important role in protein trafficking as well as have clinical significance in the release of inflammatory mediators e.g. histamine, bradykinin and cytokine release.

Purification and characterization of a human bradykinin binding protein from inflammatory cells.

Bradykinin (BK) is a potent mediator with a broad spectrum of pharmacological and inflammatory actions which are exerted through cell surface receptors. We report here the affinity chromatographic purification of a novel 14 kDa BK binding protein from human blood neutrophils and also peripheral blood mononuclear cells (PBMC), 80% of which are lymphocytes. Radioreceptor crosslinking experiments using bifunctional crosslinkers and radiolabelled BK identified a 14 kDa protein in these cell types both on the cell surface, in glycerol purified plasma membranes and in detergent solubilized cell extracts. Purification by BK affinity chromatography from a variety of BK responsive human cell types i.e. CCD-16Lu lung fibroblasts, HL60 promyelocytes, U937 myelomonocytes and Jurkat T lymphocytes also demonstrated a 14 kDa protein. Purified material obtained from three different BK affinity columns all demonstrated three major proteins at 190, 50 and 14 kDa when eluted with either excess BK or mild acid. Neutrophil fractions from detergent solubilized cell extracts contained an additional 150 kDa protein when eluted with mild acid. Neutrophil and PBMC crude plasma membrane BK affinity column purifications yielded only a single 14 kDa protein. Radioreceptor dot assays of the purified neutrophil eluates containing the 14 kDa protein revealed specific binding to [125I]-BK with a 160 fold excess signal ratio over the original membrane extract. Our data indicates that we have successfully isolated a 14 kDa novel human BK specific binding protein expressed on the surface of inflammatory cells.

High resolution autoantibody detection by optimized protein G-horseradish peroxidase immunostaining in a western blotting assay.

We report a procedure for optimisation of Western blotting using protein G-horseradish peroxidase (protein G-HRP) which avoids the false positive reactions often caused by second antibodies and increases the detection of autoantibodies by protein G conjugate. A number of modifications were investigated. Higher concentrations of serum and protein G-HRP at 1:5 to 1:10 and 1:100, respectively, increased the detection to the same order as that obtained with second antibody systems and gold staining with silver enhancement. The role of various detergents in the procedure was established. 3-[(3-Cholamidopropyl)-dimethylammonio]-1-propanesulfonate (CHAPS) in incubation with protein G-HRP increased the binding between protein G and immunoglobulin G. Addition of Tween-20 for blocking produced little background so that protein blockers could be avoided. Prolonged incubation with serum increased markedly the sensitivity of the procedure when compared with the recommended 2 h incubation period. Polyvinylidene difluoride membrane provided better transfer effect, lower background and higher mechanical strength than nitrocellulose membrane. The utilization of only one antibody-specific ligand increased the simplicity, reliability, economy, efficiency and specificity of the method. These modifications make this method significantly better for detection and screening for autoantibodies.

Coronary artery spasm and myocardial infarction in the absence of angiographically demonstrable obstructive coronary disease.

Myocardial infarction due to spasm of an "angiographically normal coronary artery" is rare, and its significance as a cause of myocardial infarction in patients without obstructive coronary disease has not been determined. Two patients are described with transmural myocardial infarction, nonobstructive coronary arteries, and suggestive evidence of coronary vasospasm as the cause of infarction. In one patient, angiography was carried out within 7 days of infarction and the documentation of normal coronary arteries argued strongly against a thromboembolic cause for infarction. This patient also had the combination of asthma, hypereosinophilia, and a systemic disease suggesting an immunologic disturbance with increased autoantibody production. A temporal association was noted between the episodes of asthma and those of coronary spasm during exacerbation of the eosinophilia, which raised the speculative issue of "allergic" coronary vasospasm. It is concluded that spasm of a nonobstructed coronary artery may cause transmural myocardial infarction. Further documentation is required in order to place this association in perspective among the other potential cause of infarction in patients with normal coronary artery anatomy.

An age-related decline in melatonin secretion is not altered by food restriction.

Melatonin has been found to exhibit youth-maintaining and disease-preventing properties. The current study examined whether the age-retarding regimen of chronic food restriction (FR) slowed the decline in melatonin secretion reported to occur with age. Total nocturnal melatonin secretion was assessed by radioimmunoassay of the primary metabolite, 6-sulphatoxymelatonin (6-S-OH-MLT), in urine. Measurements were made through adulthood (70 to 765 days) on male Wistar rats maintained on the FR regimen (60% of the normal intake) with the control animals fed ad libitum (AL). The data of animals exhibiting gross pathology were excluded. Analyses of covariance found the FR regimen had no effect on either the levels or pattern of decline observed in 6-S-OH-MLT excretion through adulthood. However, the FR body-weight-indexed metabolite measures were approximately double those of the AL (p = .06). The possibility that this result may reflect unusually high melatonin peaks in the FR tissues is discussed.

Material prepared from normal brain inhibits migration of leukocytes from Huntington's disease patients.

Material inhibiting migration of leukocytes in vitro from Huntington's disease patients, was obtained from human brain removed at autopsy. The active material was present in brainstem, basal ganglia and cerebellum but not in cerebral cortex. It could be recovered at autopsy up to 21.5 h after death, but not from patients who had died following prolonged anoxia. It was susceptible to heating and to freezing and thawing. Its activity was preserved in acid medium to pH 3, and its iso-electric point was around pH 5. Activity was present in the fraction eluted from gel filtration with molecular weight of 1.9 X 10(5). Protein, carbohydrate, lipid and nucleic acid could be identified in the active material, and its activity depended on intact protein and carbohydrate moieties. Sialic acid, although present, did not contribute to antigenicity. Extraction with lipid solvents abolished activity.

Observed differences in postoperative lymphocyte transformation are explained by patient and population variations.

In the present study, the kinetics of the lymphocyte response to phytohemagglutinin were examined by harvesting lymphocytes at 24 hour intervals in four nonoperated control subjects, preoperatively and postoperatively in six men who underwent herniorrhaphy, and in four people who had colectomy for adenocarcinoma. Nonorthogonal analysis after logarithmic transformation and adjustment for individual and population variations showed no differences in preoperative and postoperative transformation. No age effects were seen. The previous evidence that surgery inhibits transformation to phytohemagglutinin was explained by wide kinetic variations in both patient and control populations.

Delayed hypersensitivity to tetanus toxoid in man: in vivo and in vitro studies.

The frequency of positive delayed hypersensitivity skin test reactions to tetanus toxoid in 47 healthy volunteers was 80%. The frequency of immediate hypersensitivity was low (8%) and there was less discomfort than with streptokinase or PPD. There was good correlation between leukocyte migration inhibition and delayed hypersensitivity skin testing, and a quantitative relationship was demonstrated between diameter of cutaneous reaction and degree of leukocyte migration inhibition. No relationship was demonstrated between any measures of immune responsiveness and the interval from the last booster immunization. It was concluded that tetanus toxoid is a valuable antigen for assessment of delayed hypersensitivity in man.

Sensitivity tests. Their place in asthma and hayfever.

Sensitivity testing forms an essential part of the assessment of allergic disorders such as hayfever and asthma. Allergens play an important part in precipitating and perpetuating symptoms, and identification of allergens should result in significant modifications in management. Sensitivity tests demonstrate Immunoglobulin E (IgE) antibody specific for particular allergens that are suggested by the history. History, physical findings, and sensitivity tests need to be interpreted in conjunction with one another.

Diagnosing allergies. The hows and whys.

Allergies are a common problem dealt with in family practice. This article reviews the importance of a proper management approach and details the essential features in establishing the diagnosis with special emphasis given to the importance of good clinical skills.

Lymphocytes and specificity of eosinophilia.

Repeated injections of soluble antigen derived from the body fluid of Ascaris lumbricoides induced a substantial accumulation of eosinophils in the peritoneal fluid of mice. Evidence of an underlying immune mechanism was sought. A single injection of antigen was ineffective in eliciting eosinophilia, but it primed recipients to respond to a subsequent injection 30 days later. Furthermore, this priming effect could be transferred with lymphoid cells. Depletion of T cells by adult thymectomy and irradiation abrogated the ability of mice to respond. Specificity was demonstrated in primed lymphoid cells which imparted to recipients the capacity to react with increased numbers of eosinophils to a single injection of soluble antigen. Unprimed donor cells, or those primed with an unrelated antigen, were incapable of transferring the capacity for an eosinophil response. Evidence has been presented to suggest that lymphocytes which impart this specificity are T cells.

Clinical presentation and diagnosis of hereditary angio-oedema in five families.

The clinical features of 16 patients with deficiency of C1 esterase inhibitor are described. Severity ranged from no symptoms in 2 young subjects to repeated and severe abdominal pain and angio-oedema. Diagnostic techniques are described and the ease with which the diagnosis can be established is emphasized. A functional assay is required in addition to immunological determination of the protein in order to detect a minority of patients who have ineffective inhibitor. Experience of treatment with tranexamic acid is reviewed and its place in management in relation to androgens is discussed.

A study of possible ill effects from prolonged immunotherapy in treatment of allergic diseases.

It has been claimed that immunotherapy in allergic diseases may result in morbidity from chronic antigenic stimulation by the formation of immune complexes and by contributing to development of arteritis, amyloidosis and paraproteinaemia. We examined 35 older patients who received injections of allergen extracts for between two and 30 years and compared them with an age-matched control group. Treated patients had significantly higher serum IgG and lower total IgE than controls but no increased incidence of paraproteins or evidence of immune complex disease such as urinary abnormalities, increased Clq binding levels, cryoglobulins or rheumatoid fever.

Systemic lupus erythematosus in later life.

This paper presents the clinical features of 14 patients in whom systemic lupus erythematosus (SLE) was diagnosed for the first time after the age of 45 years. The onset was insidious and the diagnosis was delayed in most patients, the mean duration of symptoms before diagnosis being five years. Clinical features in this group of patients differed from classic descriptions of SLE in that there was a high incidence of neuropsychiatric disturbances and low incidence of serositis, while non-specific complaints of fever, weight loss, and malaise were often the only presenting clinical features. Factors associated with disease activity, such as elevated ESR, neutropenia, and thrombocytopenia were less frequency encountered than in younger patients. In 7 of 12 patients immunoglobulins and complement components were detected in basement membrane of normal skin. Diagnosis in this age group is difficult, and it is likely that SLE goes unrecognised in a number of older patients with non-specific complaints.

Presentation of patients with asthma at an allergy clinic in Cape Town.

A prospective study of 304 asthmatic patients attending the allergy clinic of a large teaching hospital in Cape Town was undertaken. Symptoms were overwhelmingly perennial despite a Mediterranean climate and the wealth of local flora. Allergic asthma started at any age and was frequently associated with other allergic symptoms which required treatment in their own right. Asthma in Coloured patients started later in life than in Whites and seasonal asthma was more frequent in the former group. Precipitating factors, the pattern of skin-prick test reactivity, the severity of asthma and its treatment were similar in the two groups. Serum IgE levels were higher and peripheral blood eosinophilia was more frequent in Coloureds than in Whites. Too few Black patients attended the clinic to allow for analysis.

Amyloidosis of immunoglobulin origin: useful treatment?

Six patients are presented in whom a diagnosis of amyloidosis of immunoglobulin origin was established by light and electron microscopy of biopsy samples. Treatment with a regimen of intermittent doses of melphalan and prednisone resulted in a survival of 48-82 months in four of the patients while two patients died within three months of diagnosis, too soon for the therapy to have been effective. When compared with published series which suggest that the median survival of this condition is approximately 14 months, our results suggest that this treatment prolongs life successfully. These patients also enjoyed a greatly improved quality of life while receiving treatment.