Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.3 (collagenase)
 18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We investigated the existence of an endogenous uptake system for folate in Xenopus laevis oocytes. This was done by performing uptake measurements using [3H]folic acid. Uptake of folic acid was linear with time for 4 h of incubation, and was similar in collagenase-treated and non-treated oocytes. The uptake process was carrier-mediated, as suggested by the saturation of folic acid uptake with concentration, and by the ability of unlabelled folic acid and its related compounds to significantly inhibit the uptake of [3H]folic acid. The apparent Km and Vmax of the uptake process were 42 +/- 7 nM and 10.56 +/- 0.46 fmol per oocyte per 2 h, respectively. The uptake of folic acid was independent of the presence of Na+ in the incubation medium, but was highly pH dependent with severe inhibition occurring at pH lower than 6.5. Folic acid uptake was energy- and temperature-dependent, and was significantly inhibited by the anion transport inhibitors DIDS and SITS. These results demonstrate the existence of an endogenous carrier-mediated system for folic acid uptake in Xenopus oocytes. Further characterization of the molecular mechanism of folic acid uptake and its regulation in this non mammalian in vitro unicellular system may prove useful in furthering our understanding of folate movement across biological membranes.
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PMID:An endogenous carrier-mediated uptake system for folate in oocytes of Xenopus laevis. 168 41

The duodenum, in contrast to the jejunum, actively secretes HCO3- at a high rate, a process that protects the mucosa from acid/peptic injury. Our purpose was to define the mechanisms involved in HCO3- transport by studying the acid-base transport processes in isolated duodenal enterocytes. Individual rat duodenocytes, isolated by a combination of Ca2+ chelation and collagenase, attached to a collagen matrix were loaded with the pH-sensitive fluoroprobe 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester (BCECF-AM), and intracellular pH was monitored by microfluorospectrophotometry. To identify Na(+)-H+ transport, cells in N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid 1) were pulsed with NH4Cl (40 mM) in the absence and presence of amiloride and 2) were removed of Na+. To examine Cl(-)-HCO3- exchange, Cl- was removed from Ringer-HCO3- superfusate in the presence and absence of dihydro-4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (H2DIDS). The NaHCO3 cotransporter was studied by addition and subtraction of Na+ to amiloride-treated and Cl(-)-depleted enterocytes perfused with Na(+)- and Cl(-)-free Ringer-HCO3- buffer with and without H2DIDS. Mammalian duodenocytes contain at least three acid-base transporters: an amiloride-sensitive Na(+)-H+ exchanger that extrudes acid, a DIDS-sensitive Cl(-)-HCO3- exchanger that extrudes base, and a NaHCO3 cotransporter, also DIDS sensitive, that functions as a base loader. These acid-base transporters likely play a key role in duodenal mucosal HCO3- secretion.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Proximal duodenal enterocyte transport: evidence for Na(+)-H+ and Cl(-)-HCO3- exchange and NaHCO3 cotransport. 823 51

The ability of rainbow trout liver cells to regulate their intracellular pH (pHi) was studied using two methods on hepatocytes isolated by collagenase digestion: (i) by monitoring pHi with the fluorescent dye BCECF-AM, and (ii) by measuring the amiloride-sensitive uptake of 22Na, which represents Na+/H+ exchange. In low-Na+ medium (¾16mmoll-1), Na+ uptake was reduced by approximately 70% in the presence of amiloride derivatives (DMA or MPA, 10(-4)moll-1). Changing separately either the extracellular pH (pHe) or the intracellular pH (pHi, clamped by treating the cells with nigericin in the presence of 140mmoll-1 K+) between 6 and 8 induced an increase in the rate of Na+ uptake when pHe was raised or when pHi was reduced. When transferred to hypertonic medium, hepatocytes shrank to nearly 72% of their initial volume, and thereafter a slow and partial regulatory volume increase phase was observed, with an increase in the amiloride-sensitive rate of Na+ uptake and an increase in intracellular pH. As DIDS-sensitive Cl- uptake was concomitantly enhanced, it is suggested that hypertonic stress activates Na+/H+ and Cl-/HCO3- exchange.
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PMID:Na+/H+ exchange and osmotic shrinkage in isolated trout hepatocytes 932 Feb 88

We monitored the volume of C6 glioma cells in suspension using a Coulter Counter and exposed the cells to micromolar or nanomolar levels of collagenase or clostripain. In 13 experiments, type IV collagenase (310 units ml-1; approximately 3 microM L-1) decreased the volume by 8-12%, 8 min after addition. In 13 of 21 experiments, the volume decrease was followed by a volume regulatory increase (VRI) back to control levels in the continued presence of collagenase. The shrinkage evoked by type IV collagenase was eliminated by heat-inactivation of the enzyme preparation. A highly purified collagenase (type VII) at the same concentration evoked a relatively minor decrease in volume. A well-known contaminating protease present in type IV collagenase, clostripain, which specifically cleaves arginyl peptide bonds, evoked a 7 +/- 2% shrinkage (100 nM L-1, 7 experiments). Clostripain did not evoke a volume regulatory increase. The initial velocity of shrinkage evoked by clostripain (0.0012 pL min-1, 0.0034 pL min-1, 0.0132 pL min-1; 1 pL = 10(-12) liters) scaled with its concentration (1 nM L-1, 10 nM L-1, 100 nM L-1). The effect of clostripain was inhibited by heat-inactivation of the enzyme. Leupeptin, an inhibitor of clostripain, prevented the decrease in volume evoked by clostripain. The activity of stretch-activated ion channels was unaffected by type IV collagenase. Barium, cesium, amiloride, DIDS, or bumetanide failed to block the shrinkage evoked by type IV collagenase. These results demonstrate that clostripain, present in crude collagenase enzyme preparations, causes the shrinkage, and that C6 glioma cells can undergo a volume regulatory increase at virtually constant osmotic pressure. In addition, cleavage of a cell surface moiety, which contains arginine, and possibly proline, causes shrinkage. This moiety may be part of the extracellular or intracellular matrix providing mechanical support to the cells. VRI reflect actions of another substance in the type IV crude collagenase preparations, on a receptor independent of the arg-pro moiety. The enzymatic modulation of glioma cell volume by these two receptors may reflect a new mechanism by which such cells, and possibly other glia, regulate their contact area and interactions with other cells in the central nervous system.
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PMID:Enzymatic modulation of cell volume in C6 glioma cells. 1040 29