An ultrastructural and cytochemical investigation of the development of inclusions in gastric chief cells and parietal cells of mice with the Chediak-Higashi syndrome

Gastric epithelium of the beige mouse, with a mutation thought to be analogous to that in Chediak-Higashi syndrome of man, has been examined by ultrastructural morphologic and cytochemical methods. The gastric chief cell in beige mice at 2 months of age or older disclosed two types of abnormal inclusion bodies each having distinctive morphologic and cytochemical features and a different distribution pattern and relationship to other organelles. On the basis of these findings, the first type of inclusion was thought to originate from zymogen granules, in a process of crinophagy, and the second type was interpreted as arising from the maturing face of the Golgi lamellae by the route for genesis of secondary lysosomes or lipofuscins. Each type of inclusion showed evidence both for participating in autophagic processes and for fusing with each other to produce giant inclusions. Additional observations in this study provided evidence for a role of Golgi endoplasmic reticulum lysosome in genesis of secretory granules and of the mature face of the Golgi complex in development of secondary lysosomes in chief cells. The findings also afforded evidence of migration of chief cells toward the bottom of the gland in the course of their maturation. The gastric parietal cell of control black mice disclosed secondary lysosomes, thought to arise from fusion between multivesicular bodies and mitochondria. These autophagic secondary lysosomes were enlarged in beige mice.     

The common heat shock protein receptor CD91 is up-regulated on monocytes of advanced melanoma slow progressors

Despite advances in our understanding of tumour immunology there is no therapy of proven survival benefit for advanced melanoma. Nevertheless, disease progression is slow in a small proportion of patients with metastatic melanoma, suggesting a contribution to outcome from host factors. Recent data have indicated the importance of the heat shock protein receptor CD91 in immune responses to, and progression of, infectious disease. Here we investigate the relationship between CD91 expression and outcome in malignancy. Rare melanoma patients were recruited with advanced disease that was progressing unusually slowly. CD91 expression on their monocytes was compared with control patients with more typical rapidly advancing metastatic disease. Th1 and Th2 cytokines, as well as innate and adaptive immune subsets, were also measured in the two groups. A significant increase in median CD91 expression levels was observed in slow progressors (P = 0.006). There were no differences in other immune subset markers or inflammatory cytokines. The ability of CD91 to internalize and cross-present tumour antigens through the major histocompatibility complex class I pathway may maintain CD8-positive cytotoxic T cell responses and contribute to slow progression of advanced melanoma.     

Use of Gen-Probe and Bactec for rapid isolation and identification of mycobacteria. Correlation of probe results with growth index

Gen-Probe culture confirmation tests (Gen-Probe, San Diego, CA) for Mycobacterium tuberculosis complex and Mycobacterium avium complex were performed on 276 mycobacterial isolates. All 138 M. tuberculosis complex isolates and 79 of 80 M. avium complex isolates were identified correctly. No falsely positive test results were obtained; 58 nontuberculous mycobacteria other than M. avium complex were negative by Gen-Probe. In a second phase of testing, Gen-Probe tests were performed using concentrates from 101 patient Bactec 12B cultures. Positive results by Gen-Probe tests were correlated with the growth index (GI) reading on the day of processing as well as the accumulated GI readings. For those 51 with high (greater than or equal to 999) final GIs, 40/40 (100%) M. tuberculosis complex isolates and 9/11 M. avium complex isolates were positive by Gen-Probe, and six other mycobacteria were negative. Of the 25 with moderate final readings (400 less than or equal to GI less than 999), 12/17 M. tuberculosis complex isolates and 1/1 M. avium complex isolates were correctly identified by Gen-Probe; seven other mycobacteria were negative. Of 25 with low readings (GI less than 400), 8/24 M. tuberculosis isolates were correctly identified by Gen-Probe, and no falsely positive test results were obtained with the other probes. All true negative tests on seven other mycobacteria (not M. tuberculosis complex or M. avium complex) had less than 2% hybridization. Of the 24 falsely negative tests on M. tuberculosis complex isolates or M. avium complex isolates, 22 had greater than 2% hybridization with their respective probes. Thus, percent hybridization greater than 2% may be a useful indicator of the need for retesting.     

Increased Sensitivity of Lymphocytes from Atopic Individuals to Histamine-Induced Suppression

Histamine depressed lymphocyte reactivity to phytohemagglutinin and, to a lesser degree, concanavalin A, when administered simultaneously with mitogen to lymphocyte cultures. Addition of histamine at later times to the cultures appeared to have a slightly enhancing effect on the lymphocyte response. Stimulation of lymphocytes with pokeweed mitogen was in some cases enhanced, even by high concentrations of histamine. Lymphocytes from atopic individuals were more sensitive to the inhibitory effect of histamine than lymphocytes from nonatopic individuals. The sensitivity appeared age-dependent, but within each age group histamine evoked significantly more suppression on lymphocytes from atopic than from nonatopic individuals. The possibility that the altered reactivity of lymphocytes to histamine, which appears to be associated with atopic allergy, is of pathogenic importance, is discussed, and a hypothesis for the development of atopic disease is proposed.     

Changes in leucocyte numbers without diapedesis in rats given platelet-activating factor (PAF-acether)

Changes occurring in the blood and the peritoneal cavity following the intraperitoneal injection of platelet-activating factor (PAF-acether) into rats were compared with those when antigen was injected intraperitoneally into actively sensitised rats. A blood eosinophilia had been produced in the rats by an intravenous injection of Sephadex G200 6 days before either challenge. 5 min after PAF-acether, the total number of cells in the peritoneal washings had decreased and the concentration of extravasated dye-labelled plasma protein had increased with no change in histamine levels. On the other hand, antigen at this time produced nor only a decrease in cells and an increase in dye but also an increase in histamine concentration. Only antigen produced a cellular infiltration into the peritoneal cavity with an increase in numbers of neutrophils in the peritoneal washings at 4 h and of mononuclear cells and eosinophils at 24 h. In the blood at 4 h after either challenge, there was a neutrophilia and an eosinopenia. When PAF-acether and antigen were injected together into actively sensitised rats, leucocyte counts in the peritoneal washings increased by a similar amount, both at 4 and 24 h, as those in rats given antigen alone.     

An ultrastructural study of human eosinophil granules: Maturational stages and pyroantimonate reactive cation

For ultrastructural study of the formation and maturation of human eosinophil granules, bone marrow and buffy coat specimens were fixed with an aqueous solution of potassium pyroantimonate and osmium tetroxide and by conventional methods. The antimonate-osmium tetroxide method of fixation, which is thought to permit ultrastructural localization of sodium or other cations, offered an advantage over routine methods in that it permitted recognition of four rather than two varieties of cytoplasmic granules in human eosinophils at various stages of cell development. These four granule varieties, designated A, B, C, and D, differed primarily in distribution and content of crystalloids and antimonate deposits: A granules lacked antimonate deposits and cystalloids; B granules contained a rim of deposits but lacked crystalloids; C granules possessed deposits that were present in the peripheral matrices but not in the central crystalloids; and D granules lacked deposits but contained crystalloids. Evidence is provided that these four varieties of granules represent progressive stages in the maturation of a single granule type and that granules without crystalloids are transformed into granules with crystalloids. The results also provide evidence for the presence of an as yet unidentified pyroantimonate precipitable cation in human eosinophil granules.     

The effects of drugs on leucocyte changes following the injection of antigen into the peritoneal cavities of actively sensitised rats

The injection of antigen into the peritoneal cavities of actively sensitised rats produced an immediate reaction characterised by an increase in concentrations in the peritoneal fluids, collected 5 min later, of extravasated dye labelled plasma proteins, histamine and slow reacting substance of anaphylaxis (SRS-A). Changes were also produced in the numbers of leucocytes in the blood and peritoneal cavity. 5 min after antigen challenge there was a reduction in the number of cells that could be washed from the peritoneal cavity. 4 h after antigen there was an increase in numbers of neutrophils both in the blood and peritoneal washings and these fell to the levels in control rats at 24 h. 24 h after antigen, and continuing for 72 h, there was an increase in numbers of eosinophils and mononuclear cells in the peritoneal washings.The rats were injected intravenously with sephadex particles to produce a blood eosinophilia at the time of antigen challenge, this increased the numbers of eosinophils migrating into the peritoneal cavity but had no effect on antibody levels, the numbers of other leucocytes or on the immediate reaction.An inhibitor of lipoxygenase and cyclo-oxygenase metabolism of arachidonic acid, phenidone, at 100 mg/kg p.o., inhibited SRS-A release to control levels, in the immediate reaction, but had not effect on the leucocyte changes. The glucocorticosteroid, dexamethasone, at doses of 0.1 and 1 mg/kg p.o., produced little inhibition of SRS-A release but significantly inhibited neutrophil, eosinophil and mononuclear cell infiltration into the peritoneal cavity. These results suggest that arachidonic acid metabolites released in the immediate reaction are not the prime mediators of the cellular changes. Isoprenaline, at 0.05 and 0.2 mg/kg s.c., inhibited extravasation in the immediate reaction with no effect on histamine release but only the higher dose inhibited neutrophil and eosinophil infiltration into the peritoneal cavity. Aminophylline, at 50 mg/kg p.o., had no effect on the immediate reaction but inhibited the neutrophil infiltration. Disodium cromoglycate (DSCG) at 20 and 100 mg/kg s.c. inhibited the immediate reaction but this had no effect upon the cellular changes taking place after 5 min. Cyproheptadine at 1 mg/kg s.c. inhibited extravasation but had no effect on the cellular changes. It appears therefore that factors other than those derived from the mast cell were responsible for the cellular changes in this system. DSCG at 100 mg/kg s.c. and aminophylline at 25 and 50 mg/kg p.o. prevented the reduction in the number of cells that could be washed from the peritoneal acvity 5 min after antigen challenge.     

Immunohistochemical localization of carbonic anhydrase I and II in eccrine sweat glands from control subjects and patients with cystic fibrosis.

Isozymes of carbonic anhydrase (CA) were localized immunohistochemically by the immunoglobulin-peroxidase bridge technique on fixed paraffin sections of human eccrine sweat glands. Low-activity CA I was identified in the cytoplasm of the myoepithelial cells in the secretory coil and in the luminal and basal cells of both the coiled and straight segments of the duct. High-activity CA II was found in the cytoplasm of clear cells of the secretory coil. Although evidence has suggested that CA activity is altered in cystic fibrosis (CF), the present immunohistochemical comparison of CF sweat glands revealed a distribution of and, semiquantitatively, a prevalence of CA isozymes identical to those of normal sweat glands. Abnormal enzyme activity cannot be ruled out, however, on the basis of immunocytochemical staining which depends solely on the antigenic properties of CA.     

Giant mitochondria distinct from enlarged mitochondria in secretory and ciliated cells of gerbil trachea and bronchioles

Numerous mitochondria ranging from slightly larger than normal to several micrometers in diameter (giant) were found in about one-half the serous secretory cells in the surface epithelium of the normal gerbil trachea and proximal bronchi. Tracheal serous cells of mice also were found to contain numerous giant mitochondria. Clara cells of gerbil bronchioles contained abundant giant mitochondria in addition to normal tubular mitochondria and the second population of enlarged spherical mitochondria that have been described in Clara cells of several genera. In contrast, mouse Clara cells revealed the normal tubular and the enlarged spherical mitochondria but no giant mitochondria. A survey of a number of cell types in gerbils failed to disclose hypertrophied mitochondria outside tracheobronchial surface epithelium and bronchioles. The mitochondrial enlargement resulted from an increase of matrix but not cristae. The expansion of matrix displaced the relatively sparse cristae into small collections compressed against the outer membrane. The prevalence of giant mitochondria and of granular endoplasmic reticulum is similar among cells, and these two organelles are codistributed within cells. The megamitochondria and granular reticulum occupy a central stratum, whereas normal mitochondria occur in the apical and basal regions. The giant mitochondria are considered related to a normal biologic activity that is characteristic of respiratory tract epithelium of mice and gerbils selectively and is more prominent in secretory cells than in ciliated cells.