Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A new approach, involving a two-step digestion process and Los Angeles preservation solution #1 (LAP-1), a cold storage solution, was developed for isolation of high-quality islets from human pancreata for transplantation. This approach markedly improves the islet yield, purity and viability, and the isolation success rate. In this method, the pancreas was digested first in warm collagenase solution for up to 20 minutes. After decanting the enzyme solution, partially digested tissue was dissociated by gentle agitation in cold LAP-1 solution without additional collagenase. The digested tissues were stored in cold LAP-1 solution until islet purification on Euro-Ficoll. Forty-six islet isolations were performed consecutively by the new method (group 1). These results were compared to those obtained earlier with 46 consecutive isolations, using our previous method that had been used before development of the new method (group 2). Our old method was a modification of Ricordi's method involving only warm collagenase digestion and the storage of digested tissues in cold Hanks balanced salt solution. All pancreata were partial, containing the body and tail. There were no significant differences in both groups with regard to the donor age, cold ischemic time, harvesting conditions, and pancreatic weight. Pancreas digestion was completed in approximately 1 hour in both groups. The isolation success rate as determined by viable islets after 2 days in culture was 93.5% (43 of 46 cases) in group 1, and 56.5% (26 of the 46) in group 2. Immediately after isolation, the new method yielded a total of 335,739 +/- 36,244 islets equivalent to 150 microm (IEQ) and 6,233 +/- 681 IEQ/g of pancreas with 83 +/- 2.5% purity, whereas the old method yielded a total of 195,587 +/- 25,242 IEQ and 3,763 +/- 5,509 IEQ/g with 69.2 +/- 4.7% purity. Isolated islets in group 1 maintained a good three-dimensional structure, displayed normal insulin release to high glucose stimulation in vitro, and restored euglycemia after transplantation into streptozotocin-diabetic athymic mice. The two-step digestion method provides a sufficient number of islets for transplantation from a single pancreas.
...
PMID:Improved quality and yield of islets isolated from human pancreata using a two-step digestion method. 1070 35

A molecular model of Antarctic krill euphauserase based on the known crystal structure of its fiddler crab analog, collagenase I, indicates that the core structure of these enzymes is almost identical. Euphauserase is a cold-active and thermally sensitive enzyme with a high affinity for Lys, Arg and large hydrophobic amino acids. Residue Phe137 in euphauserase, localized in loop D (autolysis loop), is highly exposed on the surface of the molecule. Therefore, it appeared to be an easy target for autolysis. The broadly specific euphauserase has a low affinity for negatively charged residues. In order to increase the stability of the enzyme, two mutants were created in which residue Phe137 was replaced by a Glu and an Asp residue. Both mutations resulted in increased stability of the recombinant euphauserase towards thermal inactivation.
...
PMID:Increasing the thermal stability of euphauserase. A cold-active and multifunctional serine protease from Antarctic krill. 1112 Nov 12

CMT-3 is a NON-ANTIMICROBIAL tetracycline (TC), chemically modified to enhance its collagenase-inhibitory property. This property is therapeutically useful in treating diseases such as periodontitis, cancer and arthritis. CMT-3 was labeled with tritium [(3)H] at Carbon 7. Four adult male Sprague-Dawley rats (350--400 g body weight) were gavaged once with a mixture of cold CMT-3 and [(3)H] CMT-3 (750 microCi). An additional four rats were gavaged for 2 days with cold CMT-3(15 mg/Kg/day) and on the third day the rats were gavaged with a mixture of cold and [(2)H] CMT-3 (750 microCi); and all 8 rats were placed in the metabolic cages. Blood samples were collected from the tail at multiple intervals from 1--14 hr after [(3)H] CMT-3 administration. At 14 hr, the rats were anesthetized, euthanized and various tissues including visceral organs were removed and weighed. The contents of GI tracts were emptied and added to the fecal pellets and weighed. The urine samples were collected and volume measured. Each tissue or organ was minced finely with scissors and 100 mg of tissue was digested in 1 ml of Tissue-solv (Packard Lab), for 4 hrs at 37 degrees C and each sample was diluted up to 10 ml of distilled water. A 100 microl aliquot was taken and diluted with an equal volume of glacial acetic acid, 10 ml of Atom-lite was added and counted for radioactivity in a liquid scintillation spectrometer. This biodistribution study revealed that over 14 hrs, 54% and 3% of [(3)H] CMT-3 were excreted in the feces and urine, respectively. The serum [(3)H] CMT-3 count reached its maximum value at about 12 hours. The tissues retained the CMTs as follow: muscle (23%); skin (2.41%); bone (1.72%); and the brain retained 0.21% of the label. The radioactive CMT-3 in the visceral organs is as follows: GI tract - its contents (8.9%); heart (0.41%), testis (0.41%); lungs >(0.16%); spleen (0.08%); liver (0.03%); kidneys > (0.02%).
...
PMID:Biodistribution of radiolabeled [(3)H] CMT-3 in rats. 1117 79

Preparation of cells from solid organs often induces a functional impairment due to the proteolytic cell damage by the applied digestive enzyme like collagenase, trypsin or dispase. To preserve the tissue and to enhance the yield of cells, Laue et al. reported an islet cell isolation with pre-incubation at 4 C permitting the enzyme to diffuse into the tissue and explicite activity equally throughout the whole particle. The aim of this study was to investigate whether this procedure can be applied to parathyroid glands. Therefore porcine parathyroid glands were dissected into 1 mm3 pieces. Subsequently one group of these pieces was incubated 22 h at 4 C in 2 mg/ml collagenase before activating the enzyme by elevating the temperature to 37 C for 30 min. The other group was incubated directly at 37 C for 30 min. The yield of cells and their viability was assessed by light-microscopy and staining with trypan-blue. After the cells were immobilized in barium-alginate and cultivated for 7 days, the function was tested by incubation in different calcium concentrations and PTH-measurement. Finally, the viability was assessed by histology. Using a cold pre-incubation with collagenase, a significantly higher number of isolated cells was retrieved compared with collagenase-digestion without pre-incubation. The viability was about 100% and did not differ between both groups. After immobilization and cultivation the viability decreased to less than 30%, with and without pre-incubation. In contrast to viability the PTH-secretion of the cells differed significantly between both procedures. By pre-incubation with collagenase at 4 C a gentle method is presented resulting in an enhancement of yield and function of single cells of parathyroid glands.
...
PMID:Improved yield and functionality of parathyroid cells separated by using collagenase-digestion with cold pre-incubation. 1126 79

For patients with chronic pancreatitis whose pain is inadequately controlled with opiate analgesia, surgical resection offers a good chance of symptomatic relief. However, the inevitable sequela is type 1 diabetes mellitus and its attendant long-term complications. Islet cell autotransplantation offers a theoretical "cure" for this iatrogenic diabetes but this end point has not been produced consistently in clinical practice. The main factor determining the likelihood of insulin independence after islet autotransplantation is the islet mass that is transplanted. This review examines the factors that affect the functional islet mass available for transplantation. Original articles and reviews from peer-reviewed journals were analyzed following a computer search of the MEDLINE database from 1966 to the present, we extracted mainly level 2 and level 3 data. Although improvements in collagenase consistency and purification techniques and reductions in cold ischemic times have all been shown to improve islet yield, there is still the need to optimize every stage in the islet isolation process. Increasing the proportion of potential islets in the final isolate is of particular importance in chronic pancreatitis because the total mass of islets initially available in the gland might be just sufficient to produce insulin independence after islet autotransplantation. We believe that reducing the warm ischemic time might significantly increase the likelihood of insulin independence after islet autotransplantation.
...
PMID:Islet yield remains a problem in islet autotransplantation. 1177 22

Drusen are abnormal extracellular matrix deposits characteristic of age-related macular degeneration (AMD), a leading cause of blindness in the aging human population. The mechanisms underlying drusen formation are not well characterized. The purpose of this study was to examine the expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in drusen, and in the surrounding cells and tissue. To assess the extent of MMP and TIMP expression by retinal pigment epithelial (RPE) cells, cDNA arrays were screened with probes generated from cultured human RPE cells. The distribution of MMP-1, -2 and -3 and TIMP-1, -2, -3 and -4 was determined using immunohistochemistry in human RPE choroid from donor eyes with and without a clinical history of AMD. Gelatinase activity was assessed in unfixed frozen sections using in situ zymography. In cultured RPE cells, expression of 10 MMP and all four known TIMP mRNAs was detected. MMP immunoreactivity was widespread in the RPE choroid, but was absent from the interior of drusen. TIMP-3, but not other TIMPs, was detected in the drusen interior. Likewise, metal ion dependent gelatinase activity could be detected in RPE choroid, but not in drusen. These results show that, while metalloproteinase activity is widespread throughout the RPE choroid, drusen are cold spots for proteolysis. The data lead to the speculation that high TIMP-3 concentrations within drusen could inhibit MMPs and as a result slow the proteolytic degradation of these deposits.
...
PMID:Drusen are Cold Spots for Proteolysis: Expression of Matrix Metalloproteinases and Their Tissue Inhibitor Proteins in Age-related Macular Degeneration. 1187 27

Two different types of brachyurins, termed I and II, have been described in the literature. Within type I there are two subtypes, Ia and Ib. The prototype for the type I brachyurins is Fiddler crab collagenase I. Its cold-adapted analogue from Antarctic krill, termed euphaulysin, shares many of its characteristics. Both enzymes are distinguished by their broad substrate specificity as well as the ability to cleave collagen. The precursor form of euphaulysin has been expressed in Pichia pastoris and processed to its fully active form using cod trypsin. A molecular model of euphaulysin, based on the known crystal structure of crab collagenase I, indicates that the core structure of these enzymes is almost identical. As a cold-adapted enzyme, euphaulysin has a higher catalytic efficiency than crab collagenase I. It is also more sensitive to thermal inactivation and autolysis. Furthermore, euphaulysin has an increased length of several surface loops compared to crab collagenase I. Extended surface loops have been suggested to play a role in the cold activity of some bacterial enzymes. Sensitivity to autolysis is an important factor which contributes to the thermal instability of euphaulysin. Substitution of a highly exposed residue in the 'autolysis loop' of euphaulysin resulted in an increased stability of the enzyme towards thermal inactivation without altering its catalytic efficiency.
...
PMID:Cold-adapted and mesophilic brachyurins. 1243 96

Low density lipoprotein (LDL) apheresis provides both structural and physiologic improvement to the vascular wall. The purpose of this study was to determine whether LDL pheresis alters levels of plasma matrix metalloproteinase-9 (MMP-9) and serum tissue inhibitor of matrix metalloproteinase-1 (TIMP-1). MMP-9 and TIMP-1 were measured in 30 healthy control subjects (Group A), 20 type 2 diabetic hemodialysis patients without obvious arteriosclerosis obliterans (ASO) (Group B), and 20 type 2 diabetic hemodialysis patients with ASO (Group C). Hemodialysis patients were dialyzed three times weekly with a bicarbonate dialysate. Twelve Group C patients underwent LDL apheresis once weekly for 10 weeks, and changes in plasma MMP-9 and serum TIMP-1 levels because of LDL apheresis were measured. LDL apheresis resulted in a significant decrease in total cholesterol and LDL cholesterol levels (p < 0.01). In addition, LDL apheresis improved clinical symptoms (including cold lower extremity, intermittent claudication, and leg pain) and diminished the size of ulcer/necrosis in all patients. Plasma MMP-9 levels were significantly higher in Group C (76.5 +/- 14.6 ng/ml) than in Group A (31.2 +/- 8.4 ng/ml, p < 0.001) or Group B (58.5 +/- 10.8 ng/ml, p < 0.05). Serum TIMP-1 levels were significantly higher in Group C (360.5 +/- 116.5 ng/ml) than in Group A (142.5 +/- 82.5 ng/ml, p < 0.001) or Group B (254.6 +/- 92.6 ng/ml, p < 0.05). Plasma MMP-9 and serum TIMP-1 levels decreased significantly after LDL apheresis (p < 0.05). However, these levels showed little change in the remaining eight Group C patients who did not undergo LDL apheresis. The data suggested that MMP-9 and TIMP-1 are associated with ASO and that LDL apheresis is effective in reducing plasma MMP-9 and TIMP-1 levels in type 2 diabetic hemodialysis patients with ASO.
...
PMID:Effects of low-density lipoprotein apheresis on plasma matrix metalloproteinase-9 and serum tissue inhibitor of metalloproteinase-1 levels in diabetic hemodialysis patients with arteriosclerosis obliterans. 1291 86

Some properties of cold neutral salt extracts of fresh guinea pig dermis have been described in terms of viscosity, electrophoresis and sedimentation patterns, partial composition, the collagen content, conditions for extraction of collagen, and the effect of certain enzymes. Viscosity of the extracts depended on the collagen in solution as demonstrated by removal of this protein by precipitation or enzymatic degradation. The intrinsic viscosity of the crude 0.45 M extract, as well as that of the isolated collagen was 14.5, identical with that for collagen dissolved by dilute acid, indicating the same high asymmetry ratio for both. Electrophoresis of the skin extracts revealed a slow moving, high, sharp, poorly diffusing boundary in addition to a pattern superficially resembling that of serum. The ultracentrifuge pattern revealed a slowly sedimenting, hypersharp boundary following a large rapidly diffusing peak. The slow moving boundaries in both patterns were abolished by collagenase or heat precipitation of the collagen fraction. Hyaluronidase had no effect on either pattern. Neutral sulfate, chloride, and phosphate extracted more collagen than did thiocyanate. Very little collagen was extracted at 37 degrees C. as compared with that removed at 3 degrees C. A two stage fractionation procedure employing dilute trichloroacetic acid and ethanol is described for the isolation and purification of soluble collagen from crude extracts.
...
PMID:Studies on the formation of collagen. I. Properties and fractionation of neutral salt extracts of normal guinea pig connective tissue. 1349 60

Normal human hepatocytes are an ideal source of liver-targeted cell therapies, such as hepatocyte transplantation and bioartificial livers, but availability of human donor livers for liver cell isolation is severely limited. To effectively utilize scarce donor organs for cell therapies, it is of extreme importance to establish an efficient isolation technique and an effective cold preservation solution for transportation of isolated cells. A lateral segment of the liver was surgically resected from pigs weighing 10 kg and a four-step collagenase and dispase digestion was conducted. Isolated hepatocytes were subjected to 8-h cold storage on ice. The following preservation solutions were tested: 1) University of Wisconsin (UW) solution, 2) UW with 100 microg/ml of ascorbic acid-2 glucoside (AA2G), 3) 100% fetal bovine serum (FBS), and 4) Dulbecco's modified Eagle's medium (DMEM) supplemented with 100% FBS. The mean viability of porcine hepatocytes was 95.5 +/- 2.5% when isolated in three independent experiments. Viability, plating efficiency, membrane stability, and ammonia metabolic capacity of cold-preserved hepatocytes were significantly better maintained by the use of UW solution. When AA2G (100 microg/ml) was combined with UW solution, such parameters were further improved. It was explained by inhibition of caspase-3 activation and retention of ATP at high levels of hepatocytes preserved with UW solution containing AA2G. The present work demonstrates that a combination of UW solution with AA2G (100 microg/ml) would be a useful cold preservation means for the development of cell therapies.
...
PMID:Maintenance of cold-preserved porcine hepatocyte function with UW solution and ascorbic acid-2 glucoside. 1457 28


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---