Crinophagic inclusions in gastric chief cells of mice with Chediak-Higashi syndrome

Immunostaining for pepsinogen with an immunoglobulin-peroxidase bridge procedure has been undertaken in conjunction with cytochemical staining for complex carbohydrate to investigate the composition and nature of large inclusions in gastric chief cells of beige mice with an analog of the human Chediak-Higashi syndrome (CHS). By these methods to stain chief cells and immunostaining for carbonic anhydrase to distinguish parietal cells, stomachs of the beige mouse were compared with those of normal black mice from which the genetic defect arose. The staining has confirmed that the chief cells are involved in CHS and has affirmed that one type of megabody in CHS chief cells contains both group I and group II pepsinogens and apparently arises in a process akin to crinophagy.     

Cytochemistry of type II pneumocytes in Chediak-Higashi syndrome of mice

The type II pneumocytes of beige mice with the Chediak-Higashi syndrome (CHS) have been compared ultrastructurally and histochemically with those of normal black mice in both newborn and adult animals. In normal black mice the inner face of the limiting membrane of lamellar bodies near the cell surface revealed continuous dialyzed iron (DI) staining as did the luminal surface of the cell. The periodate-thiocarbohydrazide-silver proteinate (PA-TCH-SP) method stained selectively the luminal surface of the granular pneumocyte together with multivesicular bodies and the inner face of the limiting membrane of lamellar bodies, in normal black mice thus supporting the role of multivesicular bodies as precursors of lamellar bodies.The beige mice lacked the layer of continuous DI staining lining the superficial lamellar bodies. Moreover, the PA-TCH-SP staining was negligible on the limiting membrane of large lamellar bodies in beige mice. Immunostaining for lysozyme appeared weaker in the large lamellar bodies of type II pneumocytes of the beige mice. Peroxisomes were normal in size but somewhat increased in number in type II pneumocytes of beige mice.The multivesicular bodies as well as the lamellar bodies increased in size and decreased in number in the beige mice and, in some cell profiles, multivesicular bodies or lamellar bodies appeared to be fusing. Since the multivesicular bodies are thought to be precursors of lamellar bodies, these findings indicate that atypical fusion of the lysosomes in type II pneumocytes occurs at early to late stages of maturation. Newborn beige mice disclosed a greater and more consistent increase in size of the lamellar bodies than did adult beige mice. Compared with newborn black mice the newborn beige mice revealed fewer secreted lamellar bodies in the alveolar space. The cytochemical and morphological changes in lamellar bodies and their precusor multivesicular bodies in type II pneumocytes of beige mice are interpreted as resulting from increased fusion of the precursor bodies and lamellar bodies with one another. Impaired secretion of lamellar bodies possibly also contributes to increased size of lamellar bodies and content of lamellar body material in granular pneumocytes of beige mice.     

Abnormally large lamellar bodies in type II pneumocytes in Chediak-Higashi syndrome in beige mice.

Lamellar bodies in type II pneumocytes are markedly enlarged in beige mice. The irregular shapes of some of these enlarged specialized lysosomes suggest fusion of smaller ellipsoidal bodies to form the giant organelles, consistent with the proposal of others for the mechanism of formation of abnormal lysosomes in Chediak-Higashi syndrome. Examination of the structure of lamellar bodies by the freeze-fracture technique reveals highly ordered stacking or concentric wrapping of the constituent lamellae. The corrugated patterns evident on the surfaces of normal lamellae are also present in Chediak-Higashi syndrome lamellae. Both normal and Chediak-Higashi syndrome lamellae appear free of typical intramembranous particles. The corrugations seen in the lamellae closely resemble those seen on freeze-fracture of dispersions of lecithin in water.     

Chediak-Higashi syndrome associated with maternal uniparental isodisomy of chromosome 1.

Chediak-Higashi syndrome (CHS) is a rare autosomal recessive disorder (incidence around 1 in 106 births), characterised by a complex immunologic defects, reduced pigmentation, and presence of giant granules in many different cell types. It most likely results from defective organellar trafficking or protein sorting. The causative gene (LYST) has recently been identified and shown to be homologous to the beige locus in the mouse. CHS has always been reported associated with premature-termination-codon mutations in both alleles of LYST. We report a unique patient with CHS, who was homozygous for a stop codon in the LYST gene on chromosome 1 and who had a normal 46,XY karyotype. The mother was found to be a carrier of the mutation, whereas the father had two normal LYST alleles. Non-paternity was excluded by the analysis of microsatellite markers from different chromosomes. The results of 13 informative microsatellite markers spanning the entire chromosome 1 revealed that the proband had a maternal isodisomy of chromosome 1 encompassing the LYST mutation. The proband's clinical presentation also confirms the absence of imprinted genes on chromosome 1.     

Heterophile antibodies in sera of patients with Chediak-Higashi syndrome

Sera of 6 patients with Chediak-Higashi syndrome and sera of their mothers were studied for heterophile antibodies. Sera of 5 patients as well as 5 sera of their mothers contained antibodies against trypsinized bovine erythrocytes, tissue sediments of guinea pig kidney or high molecular weight glycoprotein (HMWGP) of bovine erythrocyte stromata. The antibodies combining with HMWGP in enzyme immunoassay belonged to IgM and IgG classes. Although none of the sera had significant titer of agglutinins against sheep erythrocytes, on the basis of absorption and inhibition studies, these antibodies seemed to belong to the Hanganutziu-Deicher group of antibodies.     

Correction of leukocyte function in Chediak-Higashi syndrome by ascorbate.

Because ascorbate potentiates chemotaxis of normal leukocytes, we examined the effect of ascorbate on polymorphonuclear leukocytes from a patient with the Chediak-Higashi syndrome. Chemotactic migration was 104+/-16 leukocytes per 10 fields (mean+/-S.D.) initially and 258+/-44 (P less than 0.001) after ascorbate, as compared to 182+/-10 in controls. There was no bactericidal activity by 40 minutes in the patient's untreated leukocytes. After ascorbate bactericidal activity of patient and untreated control cells was the same. The addition of ascorbate reduced cAMP levels in the patient's cells from a mean of 34.5 pmoles per 10(7) polymorphonuclear leukocytes to 5.9, as compared to a control value of 3.1+/-1.4. The association of elevated cAMP and impaired function in the polymorphonuclear leukocytes of patients with the Chediak-Higashi syndrome may be related to abnormal microtubular assembly.     

Cytolytic mechanisms involved in non-MHC-restricted cytotoxicity in Chediak-Higashi syndrome.

To determine the mechanisms responsible for the impaired lymphocyte-mediated cytotoxicity in Chediak-Higashi syndrome (CHS), we investigated the killing ability of peripheral blood lymphocytes (PBL) from three patients with CHS using several kinds of target cells that were sensitive to perforin, Fas ligand (FasL), and/or tumour necrosis factor-alpha (TNF-alpha). Freshly isolated CHS PBL did not kill K562 target cells, killing of which by normal PBL was perforin-dependent, as demonstrated by complete inhibition by concanamycin A (CMA), an inhibitor of perforin-based cytotoxicity. In contrast, the CHS PBL exhibited substantial cytotoxicity against Jurkat cells, which was only partially inhibited by CMA treatment but not by the addition of neutralizing anti-FasL or anti-TNF-alpha antibodies. IL-2-activated CHS PBL exhibited substantial levels of cytotoxicity against K562 and Jurkat cells, the levels being 74% and 83% of the respective normal control values, respectively. CMA treatment showed that while the cytotoxicity of IL-2-activated CHS PBL against K562 was largely dependent on perforin, that against Jurkat was largely not. IL-2-activated CHS PBL expressed FasL mRNA, and killed Fas transfectants. These findings indicate that CHS PBL have an ability to kill some target cells via a perforin-mediated pathway, especially when they are activated by IL-2. It was also demonstrated that CHS PBL can exert cytotoxicity against certain target cells by utilizing FasL and an undefined effector molecule other than perforin, FasL, or TNF-alpha.     

Chronic active Epstein-Barr virus infection in patients with Chediak-Higashi syndrome

The results of clinical and Epstein-Barr virus (EBV) serological studies on nine Chediak-Higashi syndrome (CHS) patients are reported. Persistently elevated antibodies to the viral capsid antigen (VCA) and the restricted component of the early antigen complex (EA-R) developed in six patients who experienced primary EBV infection which either remained silent or were accompanied by clinical signs of infectious mononucleosis (IM). Hepatosplenomegaly and moderate lymphadenopathy, both clinical signs of the accelerated phase, remained detectable in the six patients for a long period of time after seroconversion. The clinical, serological, and histopathological observations are suggestive of a nonmalignant lymphoproliferative disease and consistent with an immunodeficiency to EBV. The abnormal serological responses to EBV in CHS are therefore considered manifestations of a chronic active EBV infection which may result in lethal lymphoproliferation. The three as yet seronegative CHS patients revealed no signs of the accelerated lymphoproliferative phase of the syndrome.     

Animal model. Light and electron microscopy of hepatocytes of cats with Chediak-Higashi syndrome

Cats with a condition resembling human Chediak-Higashi syndrome (CHS) are the most recently described of five species of animals with similar syndromes. In this study, hepatocytes of cats with CHS were examined by light microscopy, histochemistry, and transmission electron microscopy. Enlarged cytoplasmic granules, morphologically consistent with lysosomes, were present in many of the CHS cat hepatocytes. The enlarged lysosomes were generally larger and more numerous in centrilobular hepatocytes and were generally larger in older cats. The lesions were similar to those reported in other species with CHS suggesting that CHS cats are a valid animal model of human CHS.     

Age-related accumulation of amyloid inclusions in adrenal cortical cells.

Cytoplasmic fine fibrillar inclusions with properties of amyloid occur as neurofibrillary tangles in the brain and in the aging choroid plexus. In the present study we show that inclusions, similar but not identical to those in the choroid plexus, are common in the adrenal cortex of elderly persons. The inclusions consist of aggregates of parallel fine fibrils, often in contact with lipid droplets and partially limited by a membrane. The inclusions have affinity for Congo red and exhibit a bright green birefringence after this staining. Therefore, the inclusions can be regarded as a form of senile amyloid.     

Distribution pattern of lysosomal granules in fibroblasts of the Chediak-Higashi syndrome.

Cultured fibroblasts from a patient with the Chediak-Higashi syndrome, the mother of the patient, and a normal control were studied by light and electron microscopy. The distribution pattern of PAS-positive and acid phosphatase-containing granules in the cytoplasm differed significantly in the fibroblasts from the patient when compared with those from the mother and control. The granules in the fibroblasts from the patient were clustered in the perinuclear area, whereas the granules in the fibroblasts from the mother and control were dispersed throughout the cytoplasm. After incubation with ascorbic acid, the clustered granules in the fibroblasts of the Chediak-Higashi syndrome showed a tendency to spread throughout the cytoplasm. The distribution pattern of the granules was studied by quantitative morphology.     

Hepatic triglyceride accumulation and the ethanol physical withdrawal syndrome in mice.

Physical dependence on ethanol was induced in TO strain mice by chronic administration of ethanol by inhalation. The severity of the behavioral syndrome of withdrawal from ethanol was quantified by a subjective scoring method. During the chronic administration of ethanol, triglycerides accumulated in livers of male or female mice with a time course similar to that of the induction of physical dependence. When ethanol was withdrawn from adult or weaning dependent mice, a relationship was observed between the decline of triglyceride concentrations in liver and the duration of the ethanol withdrawal syndrome. The addition of DL-carnitine (7% w/w) to diet during the administration of ethanol markedly inhibited the accumulation of triglycerides, and significantly reduced the intensity of the ethanol withdrawal syndrome. Administration of carbon tetrachloride ((1.3 ml/kg i.p.), however, although augmenting hepatic triglyceride accumulation, had no significant effect on the withdrawal syndrome. The results are interpreted as suggesting either that ethanol-induced liver dysfunction plays a part in dependence, or, more likely, that triglyceride accumulation reflects an ethanol-induced metabolic disorder which is itself related to the induction of dependence.     

Age-related amyloid beta protein accumulation induces cellular death and macrophage activation in transgenic mice

In view of the importance of amyloid beta protein accumulation in Alzheimer's disease, this paper examines age-related amyloid beta protein (Abeta) deposition and accompanying cellular changes in a mouse model in vivo. Transgenic mice were studied which expressed a gene encoding 18 residues of signal peptide and 99 residues of the carboxyl-terminal fragment (CTF) of the Abeta precursor, under the control of the cytomegalovirus enhancer/chicken beta-actin promoter. In the pancreas, Abeta accumulated in an age-dependent manner. Abeta deposits appeared as early as 3 weeks of age and increased in size and number from 4 to 16 months of age. The largest Abeta deposits were observed in the transgenic pancreas at 16 and 20 months of age. Haematoxylin and eosin staining, macrophage immunostaining, and electron microscopy showed that the Abeta fibril deposits closely correlated with degeneration of pancreatic acinar cells and macrophage activation. Abeta1-42 and Abetap3E-42 were predominant components of Abeta deposits among amino- and carboxyl-terminal modified Abeta species. These findings suggest that overproduction of Abeta causes age-related accumulation of Abeta fibrils, with accompanying cellular degeneration and macrophage activation in vivo.     

Age-related accumulation of Reelin in amyloid-like deposits

Accumulating evidence suggest that alterations in Reelin-mediated signaling may contribute to neuronal dysfunction associated with Alzheimer's disease (AD), the most common form of senile dementia. However, limited information is available on the effect of age, the major risk factor of AD, on Reelin expression. Here, we report that normal aging in rodents and primates is accompanied by accumulation of Reelin-enriched proteinous aggregates in the hippocampal formation that are related to the loss of Reelin-expressing neurons. Both phenomena are associated with age-related memory impairments in wild-type mice. We provide evidence that normal aging involves loss of Reelin neurons, reduced production and elimination of the extracellular deposits, whereas a prenatal immune challenge or the expression of AD-causing gene products, result in earlier, higher, and more persistent levels of Reelin-positive deposits. These aggregates co-localize with non-fibrillary amyloid-plaques, potentially representing oligomeric Abeta species. Our findings suggest that elevated Reelin plaque load creates a precursor condition for senile plaque deposition and may represent a critical risk factor for sporadic AD.