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K H Sit

Publications and source records attributed to K H Sit.

At least 73 records · Page 4Linked to original sources
PubMed1990

Antiport-mediated cell retraction: viable rounding and distinctive endocytosis.

Cells that are detached through Na+/H+ antiport-mediated cell retraction (the flat-to-round or FTR change) are as viable as trypsinized cells despite unusually high trypan blue dye inclusion. Even high molecular weight dextran and carbon particles are easily loaded following the FTR change. ECM of confluent cultures of human keloid fibroblasts and Chang liver epitheloid cells stained directly by fluorescein-conjugated monoclonal anti-fibronectin is dramatically reduced or removed upon cell rounding. At the ultrastructural level, distinctive channels and vacuoles filled with extracellular material are seen, resolved at later stages as peripheral granular patches which too disappear when the rounded cells are reflattened in round-to-flat (RTF) change. Antiport-mediated endocytosis (AME) could explain the drastic reduction in cell area concomitant with cell rounding and detachment. AME provides a new way of loading cells with impermeant macromolecules.

Amiloride↗
PubMed1989

Induced interphase cell retraction: its reversal and EGF potentiation.

Human epidermoid A431 carcinoma cells retract into globular shapes upon activation with epidermal growth factor (EGF) but not normal cells. We find that normal anchor-dependent cultured cells will retract from attachment on incubation in saline with pH upshifting. There is significant potentiation of that phenomenon by EGF prepulsing as seen in (a) dose- and time-response experiments quantitated by measuring profile area and perimeter of cells (b) 22Na+-uptake studies and (c) pHi measurements. EGF action by itself produces quantitatively significant retractions, although only to a very small degree. Blocking Na+/H+ exchanges using (a) trifluoperazine (150 microM), (b) amiloride (2 mM) and (c) Na+-omission could block the cell retractions. Retracted cells can be reversed in low [Na+]o during which rapid 22Na+-efflux is observed. A dose-response ouabain (1 mM) sensitive relationship exists between [Na+]o and retardation of the reversal phenomenon. Ouabain however does not prevent reversal in low [Na+]o, nor even in combination with high [Na+]o given the presence of Ca2+. Retraction into spherical shapes would maximize volume-holding for a given surface area, however they are quantitatively more bulgy than cells rounded by trypsinization.

Amiloride↗
PubMed1989

Comparing observed average numbers of affected sibs with those expected under geometric ascertainment bias in an extended range in both simplex and multiple sibships.

Detection bias in recessive ascertainment is generally considered to be confined in a narrow range between unbiased truncate ascertainment and single ascertainment, where methods of segregation analysis are established. While there are arguments for an extended range of analysis, a deflated detection progression below the unbiased level is still being considered as theoretical ground or ignored as sporadics. I show here a method of gauging the ascertainment levels of surveyed data in a geometric continuum. The method is valid for recessive segregation at any ascertainment level and in simplex or multiplex sibships of whatever degree of truncation. Four previously published surveys are used to show conformation with real data and the existence of detection trends spanning the range from the unsuspected very depressed bias level to the inflated level above single ascertainment.

Albinism↗
PubMed1988

Kinetics of sulphate conjugation in extracts of human foetal liver cells in culture.

Sulphate conjugation was investigated using extracts of human foetal liver cells in culture. The three reactions which are involved in sulphate conjugation were measured singly or in combination: they are (i) the PAPS generation catalyzed by ATP-sulphurylase and adenosine 5'-phosphosulphate (APS)-kinase, (ii) the phenolsulphotransferase (PST) reaction, and (iii) the overall sulphate conjugation which comprises the above three reactions. All were radiometric assays employing PAP35S or sodium 35sulphate. N-acetyldopamine (NADA) was the substrate of choice although the reactions were also demonstrated with dopamine and 1-naphthol. Kinetic studies with NADA showed two pH optima of 6.7 and 8.6 for the overall sulphate conjugation and the PST reaction while the PAPS generation occurred maximally at pH 8.0. The apparent Km value for NADA measured by both the PST and the overall sulphate conjugation reactions was the same, being 38 microM, while that for inorganic sulphate, of 107 microM and 240 microM (measured by the overall sulphate conjugation reactions and by PAPS generation, respectively) was two orders of magnitude higher than that of PAPS, which was 2.57 microM. It was possible to maintain a relatively constant level of the three activities of sulphate conjugation in confluent, quiescent cultures. The importance of sulphate conjugation for detoxification in foetal cells is discussed.

Adenosine Triphosphate↗
PubMed1988

Does PAPS generation determine the overall sulfate conjugation in human platelets?

The formation of 3'-phospho-adenosine-5'phosphosulfate (PAPS) depends on essentially two enzymic reactions catalysed by ATP-sulfurylase and APS-kinase (adenosine 5'-phosphosulfate-kinase). In this paper, PAPS generation by human platelets was determined by the transfer of 35sulfate from PAP35S formed in vitro to N-acetyldopamine (NADA), using the phenolsulfotransferase extracted from rat liver. A pre-requisite of this quantitative procedure was the prior inhibition of the sulfate-activating system in the latter enzyme preparation. This was accomplished by the addition of 10 mM EDTA and 14 mM pyrophosphate. The PAPS-generating system of human platelets exhibited two pH peaks with higher activity at pH 8 than pH 6. Optimal concentrations of ATP and Mg++ at 7 mM were required for the two reactions. PAPS generation so measured showed a highly significant correlation with the overall sulfate conjugation of NADA: a correlation coefficient of 0.96 was established from data obtained from 60 platelet preparations of normal subjects.

Adenine Nucleotides↗
PubMed1988

The human platelet as an independent unit for sulfate conjugation.

The human platelets possess a full complement of enzymes capable of synthesizing N-acetyldopamine (NADA) 35sulfate from ATP, Mg++ and sodium 35sulfate. The pH optimum for this three-step overall sulfate conjugation (comprising of the ATP sulfurylase, APS kinase and phenolsulfotransferase reactions) is 8.6 and the reactions proceeded progressively for several hours. Both ATP and Mg++ ions, above their respective optimal concentrations of 5 and 7 mM, inhibited the sulfate conjugation of NADA. The apparent Km values for NADA as determined by the phenolsulfotransferase (PST) and overall reactions were similar in magnitude being 2.6 and 4.8 microM, respectively, while that for sodium 35sulfate was 202 microM. A comparison of these two activities in 62 platelet preparations of normal subjects showed that the rate of the PST reaction was generally higher than the overall reaction even though the PST assay was carried out at suboptimal concentration of PAPS. There was a positive correlation (r = 0.82) between the two sets of data, suggesting that the PST reaction probably has some control over the rate of overall sulfate conjugation.

Adenosine Triphosphate↗
PubMed1980

Three levels of G-band differentiation.

G-band "fusions" in the more contracted chromosomes, and G-band "splittings" in the longer chromosomes, have been cited directly or by implication to be natural cyclical states of human chromosomes. This report presents three levels of G-band differentiation producted by a single techinque, and compares them with three corresponding levels of R-band differentiation reported by others. It argues for the possibility of artifactural band "fusion" mechanisms in the more contracted chromosomes.

Azure Stains↗
PubMedDate not supplied

Cytochrome P450 content and ultrastructural changes in liver of zinc-treated C57/6J mice.

In a previous report, we have demonstrated that acute zinc administration reduced the hepatic cytochrome P450 content in female C57/6J mice. In this extended toxicological study, we investigated the effects of zinc administration on (a) the hepatic cytochrome P450 content of both male and female mice to evaluate whether the sex of the animal had any influence on the zinc effects and (b) the hepatocytes at the ultrastructural level. Two doses of zinc chloride at 28 micrograms g-1 body weight (equivalent to LD50 in chronic treatment) were administered intraperitoneally to male and female C57/6J mice at 24 h intervals. Significant reduction of hepatic cytochrome P450 content was observed to occur the next day in both acutely treated male and female mice. On examination under transmission electron microscopy, evidence was found of toxic injury to the hepatocytes of mice livers in the zinc-treated group. Glutathione-monochlorobimane adduct formation (which is specifically catalysed by glutathione transferase) was found to be depressed in Chang liver cells. The findings indicate that acute zinc administration reduced the hepatic cytochrome P450 content in C57/6J mice irrespective of gender.

Animals↗