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K H Sit

Publications and source records attributed to K H Sit.

At least 37 records · Page 2Linked to original sources
PubMed1996

Effect of zinc on the epithelial lining of mice epididymis--a light microscopic study.

Intraperitoneal zinc chloride was administered at 7.5 micrograms/g body weight and 15 micrograms/g body weight to 10-12 weeks old Swiss albino mice for 5 consecutive days. Control animals were given normal saline. The testis and epididymis were dissected and examined under the light microscope. Micrographs of the testes appeared normal in both treated and nontreated animals. However the group of animals treated with the higher dosage of zinc chloride showed evidence of rupture and collapse of the epididymal epithelial lining. The testes were not affected probably because of (a) known higher testicular concentration of metallothioneins which can bind the zinc and consequently detoxify the metal and (b) "stratified" epithelium comprising of spermatogenic and Sertoli cells.

Animals↗
PubMed1996

Acidification and recovery results in nuclear accumulation of supravital dyes during interphase.

Recent studies using real time imaging demonstrated relative nuclear insulation for ion-size particles. We show here that acidification and recovery converted the insulated interphase nuclei of KB carcinoma and nontumorigenic Chang cells into intense nuclear accumulating states marked by sequestration of the exogenous supravital dyes neutral red, methylene blue, and brilliant cresyl blue. The phenomenon was not affected by Na(+)-free and HCO3(-)-free conditions nor by the presence of cationic and anionic antiport regulators of cytosolic pH. Cytological, microspectrophotometric, and flow cytometric evaluation of whole cell populations showed that the nuclear influx was abolished by omitting the pH recovery response, and by modulating the recovery response. The abolition of nuclear influx in the presence of the P-ATPase and Fzero-ATPase inhibitors, vanadyl(IV) ions and oligomycin, respectively, suggest that H(+)-translocating ATPase pumps are involved in regulating cytosolic acidification in Na(+)-free and HCO3-conditions vanadyl(IV) inhibited nuclear uptake of supravital dyes in a dose dependent manner. Nuclear uptake of dyes, however, was not affected by up to 1 mM of genistein even though tyrosine-specific phosphorylation and DNA synthesis were abolished. Upgradient nuclear influx involving proton pump is novel. KB cancer cells and nontumorigenic Chang cells had differential dye accumulations induced by acidification and recovery.

Cell Line↗
PubMed1996

Cell rounding with "rip off" detachment.

The "rounding up" characteristic of mitotic, apoptotic, transformed and non-transformed cells appears to be initiated by various causative factors. Cytoskeletal and adhesion modulations have been implicated. Endocytic internalization via large channels seen in the rounding of amoeba and human cells alike, implicate a third mechanism, viz. membrane flow and fusion, where "rip off" detachment sheds parts of the cell to achieve freedom. Mutilation as a means for detachment is also used by moving fibroblasts, linking shape change with movement via a distinct mechanism. While rounding may follow detachment and detachment may follow rounding, "rip off" mutilation, whether self-inflicted or imposed, and modulation of stickiness are fundamentally different processes.

Animals↗
PubMed1995

Acute zinc administration prolongs hexobarbitone-induced sleeping time in C57/6J mice.

Two experiments were carried out in this study to demonstrate the effects of acute intraperitoneal (i.p.) administration of zinc chloride (ZnCl2) on hexobarbitone-induced sleeping time (HST) in female C57/6J mice. Serum and hepatic zinc content as well as hepatic cytochrome P450 content were also estimated in ZnCl2-treated and control mice. The ZnCl2 dose used was equal to the LD50 for chronic treatment (which was 28 micrograms g-1 body weight for mice given i.p. ZnCl2 five times a week for 3 weeks), determined in an earlier experiment. ZnCl2 injections at this dose were either given singly (in experiment I) or repeated on two mornings (in experiment II). Hexobarbitone, an ultra-short-acting experimental barbiturate, was given intraperitoneally 30 min after the single ZnCl2 injection in experiment I and 24 h after the second ZnCl2 injection in experiment II. Appropriate controls were given i.p. normal saline in each experiment. The HST was observed for all the animals, using the time-points at which the loss and regain of righting reflex occurred as the parameters. The animals were later killed; their blood and livers were obtained for estimation of zinc levels and cytochrome P450 content. The results of both experiments showed that ZnCl2 had caused a significant prolongation of the HST in C57/6J mice. Serum and hepatic zinc content were also elevated in both groups of ZnCl2-treated mice compared to their respective controls. The cytochrome P450 content in the single-dose ZnCl2-treated mice was unchanged while it was significantly reduced in the double-dose ZnCl2-treated mice when compared to the content in their respective controls. These findings suggest that acute zinc excess has an inhibitory effect on the function as well as the synthesis of hepatic cytochrome P450 enzymes.

Animals↗
PubMed1995

Effect of intraperitoneal zinc on the haematological profiles of C57/6J mice.

Zinc is included in a number of medications and haematotoxic effects due to zinc excess have been reported. In this study, the haematological profiles (haematocrit, haemoglobin, total white blood cell count, differential white blood cell count and platelets) of mice treated with zinc were evaluated. Intraperitoneal zinc chloride was administered to C57/6J mice in varying dosages from 1.4-14 micrograms/g body weight, four times a week for a period of three weeks. Zinc chloride had no effect on the haematological profiles of these mice since the blood cell counts of treated mice were not significantly different from the controls (p < 0.05), with the exception that at half LD50 of zinc chloride (14 micrograms/g body weight) a reactive thrombocytosis resulted. (The platelet counts between the control and experimental group of mice were significantly different, exceeding 95% confidence limits; p = 0.016). We postulate that the specific effect on platelets was due to zinc being a potent inhibitor of phenol sulfotransferase (PST), an enzyme which is involved in many metabolic pathways. Platelets are a rich source of PST and the thrombocytosis observed was probably a compensatory mechanism to raise the levels of PST in the body.

Animals↗
PubMed1994

Reduced surface area in apoptotic rounding of human Chang liver cells from serum deprivation.

BACKGROUND: The early stages of apoptosis (programmed cell death) are said to be characterized by internucleosomal DNA fragmentation and "condensation of the cytoplasm" in which cells round up, detach, and increase in density. We studied the causation of apoptotic rounding. METHODS: Human Chang liver cells in normal monolayer culture were compared with apoptotic counterparts derived from serum growth factor deprivation. Cell-by-cell analysis using the Coulter EPICS PROFILE II flow cytometer studied 1) the cell cycle from propidium iodide-DNA bindings, 2) uptake of neutral red (NR) dye, a viable cell marker, and 3) cytosolic pH (pHi) modulations from 2',7'-bis(2-carboxyethyl)-5(and-6)-carboxyfluorescein (BCECF) fluorescence ratios with NH4Cl prepulsing and forward scatter bitmapping of cell surface area. Morphometric studies were done in the Quantimet 570 image analyser. Uptake of trypan blue, neutral red, and 2 million mol.wt fluoresceinated dextrans was studied by light microscopy. Cytological profiles were examined in light microscopy and transmission and scanning electron microscopy. RESULTS: Three days of serum growth factor deprivation caused confluent flat substrate-attached cells to retract and round up, tethering tenuously to the substrate via thin microvillus attachments only. Ninety percent of cell surface area was lost with this flat-to-round change. There was high trypan blue staining with total loss of proliferative potential, and the entire genome was just fragmented DNA making up the solitary Ao (apoptotic) peak in cell cycle profiles. However, these rounded apoptotic cells also internalized huge 2 million mol.wt dextran particles and impermeant neutral red which is an established viable cell marker. The rounded apoptotic cells had an intensely acidic (pH 5.6) cytosol and therefore a steep [H+]i/[H+]o gradient promoting proton extrusion. The pHi upshifted dynamically upon acidification, recovering and even exceeding resting level by a whole pH unit. Surface area reduction occurred concomitantly in real time with pHi upshifts in these apoptotic cells. Acidification and recovery in apoptotic cells also produced enhanced uptake of neutral red. Cytological profiles showed abundant large endocytic channels and endosomes in the rounded apoptotic cells. CONCLUSIONS: Gross surface area reduction with evidence of distinctive endocytic activity including uptake of huge 2 million mol.wt dextran particles suggested large channel endocytic internalization as a causal factor in apoptotic rounding, in common with rounding in M-phase and interphase cells with pHi upshifting where concomitant surface area reduction and uptake of impermeant particles were similarly demonstrable. The reduction in size of the cell envelope, together with consequential concentration pressures, could account for the observed rise in cell density and shrinkage in cell size. As a symptom of continual pHi upshifting, apoptotic rounding appears to be a recovery-associated response rather than a direct consequence of the disruptive forces causing its death.

Apoptosis↗
PubMed1994

Lymphocytic infiltration in focal thyroiditis: an ultrastructural case study.

In autoimmune thyroiditis, infiltrating lymphocytes play an important role in the disease process since they are known to release cytokines which can alter thyroid functions. The degree and site of lymphocytic infiltration in the thyroid gland have been used to differentiate between Hashimoto's disease and focal lymphocytic thyroiditis. Intraluminal lymphocytes have been reported to be absent in focal thyroiditis but present in Hashimoto's disease. However, we demonstrate here by transmission electron microscopy, that extrafollicular and intrafollicular lymphocytes could be present in focal lymphocytic thyroiditis. This seems to support the notion that focal lymphocytic thyroiditis could represent part of the spectrum of autoimmune thyroiditis and Hashimoto's thyroiditis.

Adult↗
PubMed1994

Azide- and vanadate-sensitive M-phase alkalinity and cytosolic acidification of Chang liver cells.

Flow cytometric cell-by-cell evaluation of NH4Cl acidification of human Chang cells showed that at steady state, 3% of the cells remained alkalinized (> pHi 7) over an extended period (up to 80 min) despite the absence of extracellular Na+ and HCO3-. In fluorescence microscopy, the acidification-resistant cells were characteristically rounded M-phase cells. Both mean cytosolic pH and M-phase alkalinity were however sensitive to (a) azide and oligomycin, inhibitors of F-ATPase (ATP synthase), and to (b) vanadium ions, the phosphate analogue of P-ATPase (ATP-hydrolyzing), in dose-dependent and time-dependent manners. Dead cell indices were constant at approximately 10%. Thiocyanate chaotrophic anions, which cleave the V-ATPase structure, had no effect. Since ATP synthesizing F-ATPase (ATP synthase) is coupled to ATP-hydrolyzing P-ATPase as 'master-&-slave', azide- and oligomycin-sensitivity corroborated with vanadate-sensitivity in suggesting energized proton pumping modulating (a) M-phase alkalinity and (b) cytosolic pH, against acidification.

Adenosine Triphosphatases↗
PubMed1994

Internalization of large macromolecules in human Chang liver cells by extracellular ATP.

The home of all intermediary metabolism and the cytoskeleton, viz. the cytosol, appears inaccessible to large macromolecules in the living state without severe perturbations. ATP-induced cytosolic penetration due to poration which allow passage of < 1000 mol. wt. particles have been cited. We report here that at similar milli-molar dosage levels of ATP, internalization of 2,000,000 mol. wt dextran particles as well as 0.92 micron and 2.17 microns latex beads could be demonstrated. At ultrastructural level, the internalized beads were seen distributed in 2 sites on the cytoplasm of the cell, viz. (a) inside vacuoles and (b) outside vacuoles, in the cytosol. The vacuolar distribution suggests endocytic entry, while the cytosolic distribution suggests a cellular sorting process where the boundary of the bead is processed or sorted out of the membrane-recycling pathway.

Adenosine Triphosphate↗
PubMed1993

Reduced surface area in mitotic rounding of human Chang liver cells.

The rounding up of mitotic human Chang liver cells in monolayer culture was studied quantitatively. It was surprising to find significant reduction in cell surface area considering that endocytosis has been demonstrated to be at a complete standstill in M phase. Uptake studies using impermeant BCECF (2',7'-bis(2-carboxyethyl)-4(5)-carboxyfluorescein free acid) pH indicator and particulate neutral red dye in aqueous buffer showed preferential internalization into mitotic cells in direct contrast to expectation since interphase cells do not have arrested endocytosis. However, infolded plasma membrane ruffles and internalized extracellular material were demonstrated in prophase cells, much like those seen in interphase rounding via the induction of intracellular alkalinizations. Raised intracellular pH (pHi) is a universal and consistent finding in M phase cells. Despite cessation of small pit endocytosis, it remains possible for plasma membrane internalization to be a causal factor in the observed surface area reduction in mitotic rounding.

Cell Membrane↗
PubMed1993

Effect of genistein, a tyrosine-specific protein kinase inhibitor, on cell rounding by pH upshifting.

Cell rounding is generally regarded as a cytoskeletal change exemplified by the mitotic state of monolayer cell cultures in which mitosis-specific phosphorylation was recently cited as the molecular mechanism. Mechanistically, there appears a convergence with earlier suggestions of tyrosyl phosphorylation of cytoskeletal elements in growth-factor-induced rounding inasmuch as protein tyrosine kinase plays a major regulatory role in cell cycle progression. On the other hand, despite the permissive association between cell activation and intracellular alkalinization, inducing rounding via pH upshifts in the presence of even high concentrations (135 micrograms/ml) of a tyrosine specific protein kinase inhibitor, genistein, did not suppress the rounding response, although a distinct difference in phosphotyrosine level was demonstrated by monoclonal antiphosphotyrosine antibodies.

Cell Line↗
PubMed1993

Cell shape changes induced by sulphate in the Cloudman mouse melanoma cell line.

The mechanism of cell shape changes and haematogenous translocations (metastases) in mouse malignant melanoma cells induced by phorbol esters and protein kinase C (PKC) is reported as adhesion "downregulation", exocytosis and motility. However, PKC activation also produces intracellular alkalinization, a causal factor in plasma membrane internalization, cell rounding and detachment that does not necessarily implicate specific cell adhesion downregulation. We show here that Cloudman mouse malignant melanoma cells can be induced to round up and detach with concomitant intracellular alkalinization by simple inorganic sulphate treatment, thereby suggesting an alternative explanation to the reported phenomena.

Animals↗
PubMed1993

Oncocytes of the thyroid: a neutral red dye uptake study.

Oncocytosis and oncocytomas have attracted attention because on the one hand there have been reports of the favourable prognosis and generally benign nature of oncocytic tumours, while on the other hand, Hurthle cell change in thyroid neoplasms have been reported to be correlated with aggressive biological behaviour and a significantly worse clinical course. Presently, there appear to be two schools of thought on the significance of oncocytosis: (i) a process where there is redifferentiation of cells with involvement in the pathogenesis of cancer; and (ii) a compensatory process for 'functional exhaustion' of the cell. Many aspects of oncocytosis seem unclear. In this study, transmission electron microscopy is used to demonstrate the presence of thyroid oncocytes in the hemithyroidectomy specimen of a patient with focal lymphocytic thyroiditis. Tissue sections from this specimen did not display preferential uptake of neutral red dye, a distinctive characteristic of M-phase cancer cells which was previously reported. The findings seem to suggest that oncocytes are not cells with distinctive proliferative potentials. If that were the case, then oncogenesis or cancer cell progression and oncocytosis may not be convergent processes.

Adenoma↗
PubMed1993

Effect of intraperitoneal administration of zinc in C57/6J mice.

Intraperitoneal administration of zinc chloride to C57/6J mice in vivo at 28 micrograms/g body weight was more lethal to the male mice and induced growth retardation in the male and female survivors. In the group of experimental survivors, the weight of the lungs were significantly decreased compared with the control animals. The inherent dangers of zinc excess is highlighted, especially with regard to the possibility of cumulative zinc toxicity.

Animals↗
PubMed1992

Human Chang liver cells show large surface openings and endocytic channels that resemble those in the amoeba.

Cytolysis of host cells by pathogenic Entamoeba histolytica can be blocked by specific lysozyme inhibitions and is recently reported to be enhanced by phosphoinositide (PI) signal transduction activation. However the mechanistic relationship between PI second messenger targets and massive lysosomal secretion needed to achieve rapid host cell lysis is unclear. We have previously shown that intracellular alkalinization associated with activated PI hydrolysis produces a massive endocytosis of huge proportions which would force a corresponding exocytosis for the maintenance of overall cell dimensions. These endosomes are processed by primary lysosomes. Apparently then, the massive exocytosis secretory pathway could provide the means for the ejection of lysozymes over target cells. We show here using human Chang liver cells that intracellular alkalinization produced large surface pittings similar to those seen in pathogenic E. histolytica in a rounded state. The SEM profile is correlated with the TEM profile of large endosomes containing extracellular debris and endosomes associated with primary lysosomal vesicles, which could support the notion that some of the pittings seen in the rounded Chang cells and the pathogenic amoebae are exit portals for endosome-lysosomes.

Animals↗
PubMed1992

Degranulation of rough endoplasmic reticulum in M phase and adenosine-triphosphate-treated interphase cells is reversible.

In the preferential harvesting of rounded mitotic (M phase) cells of human Chang liver monolayer cultures by mechanical agitation in Ca(2+)-free phosphate-buffered saline, degranulation of endoplasmic reticulum (ER) was observed. Mitotic cells are known to have a series of Ca2+ transients and, without being subjected to Ca(2+)-free washings, did not have degranulated ER. Quiescent cells incubated with 0.7 mM adenosine 5'-triphosphate (ATP) in Ca(2+)-free HEPES-buffered saline produced very similar ER degranulations. Confocal argon laser imaging of fluo-3-loaded cells showed a Ca2+ transient peaking at 2 min after ATP treatment. In the absence of extracellular Ca2+, transients of Ca2+ elevation in the cytosol would exit the cell in a down-gradient, draining the ER Ca2+ stores. Substituting ATP with 1 microM brominated A23187 calcium ionophore in the incubation that contained 1-100 mM CaCl2, respectively, did not produce ER degranulation, thereby excluding raised cytosolic Ca2+ per se as the cause of ER degranulation. In fact, incubation with 0.7 mM ATP in the presence of 1-5 mM CaCl2 failed to produce ER degranulation. ER degranulated cells, from treatment with ATP without extracellular Ca2+ as well as from Ca(2+)-free washings at M phase, could be rescued by subsequent incubation in growth medium that contains Ca2+ whereupon the rounded cells re-flatten (a round-to-flat change) and have well-defined rough ER. It therefore seems possible for Ca2+ depletion, or at least a reduction, to be causally related to ER degranulation. If that were the case, ER granularity would appear to be a facultative rather than a constitutive state.

Adenosine Triphosphate↗