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Biomedical subjects

J Chao

Publications and source records attributed to J Chao.

At least 91 records · Page 5Linked to original sources
PubMed1997

The effect of bradykinin on secretion of insulin, glucagon, and somatostatin from the perfused rat pancreas.

To evaluate the effect of bradykinin (BK) on rat islet alpha, beta, and delta cells, the rat pancreas was perfused in situ with BK (1 mumol/L) for 30 minutes via a cannula placed in the celiac artery. Insulin, glucagon, and somatostatin concentrations in the effluent were measured to determine the effect of BK on the secretion of these hormones. The BK concentration of the rat pancreas was also measured. Basal secretion of insulin, glucagon, and somatostatin in medium containing 6 mmol/L glucose was maintained at 6.5 +/- 0.5 ng/mL 124 +/- 8 pg/mL, and 511 +/- 22 pg/mL (n = 12), respectively. BK (1 mumol/L) induced a transient peak that was 3.7-fold of the baseline concentration within 3 minutes, followed by a sustained level that was approximately 50% higher than baseline. BK also transiently increased glucagon secretion with a peak that was 1.7-fold of the baseline concentration within 3 minutes, without a sustained secretion phase. BK caused a reduction in somatostatin secretion within 3 minutes to a level of 60% to 70% of the baseline concentration. The BK concentration of the rat pancreas was 3.42 +/- 1.45 micrograms/g protein (n = 5), which was approximately 3 mumol/L. We concluded that BK stimulated insulin secretion, transiently increased glucagon secretion, and decreased somatostatin secretion during the 30-minute perfusion of the rat pancreas.

Analysis of Variance↗
PubMed1997

Expression of human tissue kallikrein in rat salivary glands and its secretion into circulation following adenovirus-mediated gene transfer.

Replication-deficient adenovirus Ad.CMV-cHK, expressing human tissue kallikrein under the control of the cytomegalovirus enhancer/promoter, was introduced into rat salivary glands via a direct intracapsular injection. A single injection of Ad.CMV-cHK at a dose of 4 x 10(9) pfu resulted in a sustained expression of human tissue kallikrein in rat salivary glands. The level of immunoreactive human tissue kallikrein in rat sera was the highest at 1 day post gene delivery when both salivary glands were injected and decreased in a time-dependent manner after gene delivery. Human tissue kallikrein levels in sera increased concomitantly with the amount of adenovirus used in direct salivary injection. The detection of human tissue kallikrein in sera after gene delivery into salivary glands provided direct evidence indicating that rat salivary glands secrete locally synthesized human tissue kallikrein to the systemic circulation. The direct injection of salivary glands with replication-deficient adenovirus could provide a systemic route for gene delivery for studying salivary gland function and development. Targeted gene delivery to the salivary gland may provide the means to express therapeutic proteins in saliva and the systemic circulation.

Adenoviridae↗
PubMed1997

Kallikrein gene therapy: a new strategy for hypertensive diseases.

The tissue kallikrein-kinin system has been postulated to play a role in blood pressure homeostasis and the pathogenesis of clinical hypertension. To demonstrate the potential therapeutic effects of somatic gene delivery in treating hypertension, we used spontaneously hypertensive rats (SHR) as a model. The gene encoding the human tissue kallikrein was used because of its powerful hypotensive action. The human kallikrein DNA constructs were placed under the control of the metallothionein metal response element, the cytomegalovirus promoter/enhancer or the Rous sarcoma virus 3'-LTR. The human tissue kallikrein DNA constructs were incorporated into adenoviral vectors via homologous recombination. The naked plasmid DNA constructs or adenovirus containing the kallikrein gene were first introduced into kidney 293 cells and the expression of human tissue kallikrein was identified by ELISA. The kallikrein gene was delivered into SHR via intramuscular, intravenous, portal vein, intraperitoneal, and intracerebroventricular routes. A single injection of naked human kallikrein DNA constructs caused a prolonged reduction of high blood pressure for up to 8 weeks. Adenoviral-mediated gene delivery results in high efficiency of human tissue kallikrein expression. Immunoreactive human kallikrein was detected in rat serum at the highest level at 1 day post gene delivery. Portal vein delivery of a reporter gene, AdCMV-LacZ, results in intense staining of beta-galactosidase in rat liver, suggesting that recombinant kallikrein is mainly produced in liver and secreted into the circulation. These results show that kallikrein gene delivery causes a sustained reduction of blood pressure in genetically hypertensive rats and provide important information for a potential gene therapy approach to human hypertension and related diseases.

Animals↗
PubMed1997

Purification, characterization and activation of fish muscle prokallikrein.

Fish prokallikrein was isolated and characterized from skeletal muscle of the black sea bass, Centropristis striata. The prokallikrein was purified to apparent homogeneity by anion exchange perfusion chromatography and reversed phase high performance liquid chromatography. Initial identification was by its weak immunoreactivity with human tissue kallikrein antiserum. Two-dimensional gel electrophoresis and immunoblotting identified the protein as 36 kDa with a pI of 4.95-5.15. The prokallikrein was trypsin-activated to produce an approximately 36 kDa active enzyme as identified on an SDS-polyacrylamide gel overlayed with a membrane impregnated with the fluorogenic tripeptidyl substrate D-Val-Leu-Arg-7-amino-4-trifluoromethyl-coumarin. A potential dimer at 72 kDa was also enzymatically active. Bass kallikrein cleaved low molecular weight dog kininogen to release kinin peptide as determined by radioimmunoassay. The enzyme's amidolytic activity, with a pH optimum at 9.0, was inhibited by aprotinin, benzamidine, and phenylmethanesulphonyl fluoride, but not by elastatinal, soybean trypsin inhibitor, or limabean trypsin inhibitor. Polyclonal antiserum raised against the purified bass muscle prokallikrein recognized 36 kDa and 72 kDa proteins in bass heart, skeletal muscle, spleen, swimbladder, gill, and kidney by Western blot analyses. The wide distribution of immunoreactive proteins in the tissues suggests a potential physiological role for fish kallikreins in muscle contraction and/or relaxation, the regulation of local blood flow, and in osmoregulation. The detection of fish prokallikrein and its activation leads the way for an evaluation of the impact of kallikreins in fish health and disease processes and for studying the evolution of kallikreins and related serine proteinases.

Animals↗
PubMed1997

Distribution of tissue kallikreins in lower vertebrates: potential physiological roles for fish kallikreins.

Fish skeletal muscle prokallikrein was purified from black sea bass, Centropritis striata, and used for the production of polyclonal antiserum. Tissue proteins from primitive fish and teleosts, an alligator, and an insectivore were resolved by sodium dodecylsulfate-polyacrylamide gel electrophoresis, Western blotted, and probed with fish muscle prokallikrein antiserum. A recurring theme was the presence of approximately 36 and 72 kDa kallikrein-like proteins in skeletal muscle, heart, gill, kidney, and spleen of higher teleosts and in selected tissues of sturgeon, shark, alligator, and mole. The presence of immunoreactive kallikreins in osmoregulatory organs such as the gills of teleosts and the rectal gland of sharks signifies a potential role for these proteins in osmoregulation. Black sea bass, rock bass, and sturgeon contained many immunoreactive kallikreins in their swimbladders, which implicates a role for kallikreins in the regulation of blood flow and vascular permeability to facilitate gas exchange within the bladder. Kallikreins were consistently identified in skeletal muscle and heart of all the species evaluated and may regulate local blood flow, muscle contraction or relaxation, or participate in various transport processes. The antiserum to fish prokallikrein recognized immunoreactive kallikreins from pancreatic tissues from fish and lower vertebrates, but not from the pyloric caecum of sea bass. The wide distribution of tissue kallikrein in lower vertebrates suggests that it may participate in a variety of physiological functions.

Alligators and Crocodiles↗
PubMed1997

Regulation of bradykinin B2-receptor expression by oestrogen.

1. Tissue kallikrein is overexpressed in the kidney of female rats, this sexual dimorphism being associated with a greater effect of early blockade of bradykinin B2-receptors on female blood pressure phenotype. We evaluated the effect of ovariectomy and oestradiol benzoate (50 micrograms kg-1 every two days for two weeks) on the vasodepressor response to intra-arterial injection of bradykinin (150-900 ng kg-1) and on the expression of bradykinin B2-receptors. 2. Ovariectomy reduced the magnitude of the vasodepressor response to bradykinin and unmasked a secondary vasopressor effect. Oestrogen replacement restored the vasodepressor response to bradykinin in ovariectomized rats. 3. The vasodepressor responses to sodium nitroprusside (3-18 micrograms kg-1), acetylcholine (30-600 ng kg-1), desArg9-bradykinin (150-900 ng kg-1) or prostaglandin E2 (30-600 ng kg-1) were significantly reduced by ovariectomy. Oestrogen restored to normal the responses to desArg9-bradykinin, acetylcholine and prostaglandin E2, but not that to sodium nitroprusside. 4. B2-receptor mRNA levels were decreased by ovariectomy in the aorta and kidney and they were restored to normal levels by oestrogen. Neither ovariectomy nor oestradiol affected receptor expression in the heart and uterus. 5. These results indicate that oestrogen regulates B2-receptor gene expression and function. Since kinins exert a cardiovascular protective action, reduction in their vasodilator activity after menopause might contribute to the increased risk of pathological cardiovascular events. Conversely, the cardioprotective effects of oestrogen replacement might be, at least in part, mediated by activation of the kallikrein-kinin system.

Animals↗
PubMed1997

N1-dansyl-spermine and N1-(n-octanesulfonyl)-spermine, novel glutamate receptor antagonists: block and permeation of N-methyl-D-aspartate receptors.

The effects of several N-sulfonyl-polyamines, including N1-dansyl-spermine (N1-DnsSpm) and N1-(n-octanesulfonyl)-spermine (N1-OsSpm), were studied at recombinant N-methyl-D-aspartate (NMDA) receptors expressed in Xenopus laevis oocytes. N1-DnsSpm and N1-OsSpm inhibited NMDA receptors and were approximately 1000-fold more potent than spermine in oocytes voltage-clamped at -70 mV. Block by N1-DnsSpm and N1-OsSpm was strongly voltage dependent, being more pronounced at hyperpolarized membrane potentials. With the Woodhull model of voltage-dependent channel block, the values of Kd(0) were 779 microM, 882 microM, and 7.4 mM and those of z delta were 2.58, 2.57, and 1.07 for N1-DnsSpm, N1-OsSpm, and spermine, respectively. This suggests that an increase in the voltage dependence of block together with an increase in affinity contributes to the increased potencies of N1-DnsSpm and N1-OsSpm compared with spermine. Sensitivity to N1-DnsSpm was reduced by mutation NR1(N616Q) and was increased by mutations NR1(N616G) and NR2A(N615G). The NR1(N616G) and NR2A(N615G) mutations decreased the Kd(0) value of N1-DnsSpm without affecting z delta, whereas the NR1(N616Q) mutation reduced z delta. These mutations may alter the accessibility of part of the polyamine binding site within the channel pore or directly alter the properties of that site. Block by N1-DnsSpm (0.3 microM) was almost complete at -100 mV, and there was no relief of block at extreme negative membrane potentials (-100 to -200 mV) at wild-type NR1/NR2A channels. In contrast, block by N1-DnsSpm was partially relieved at extreme negative potentials at receptors containing NR1(N616G) or NR2A(N615G), suggesting that N1-DnsSpm can permeate these mutant channels but not wild-type NR1/NR2A channels. This is hypothesized to be due to an increase in the pore size of channels containing NR1(N616G) or NR2A(N615G), which allows passage of the bulky head group of N1-DnsSpm. In contrast to N1-DnsSpm, N1-OsSpm could easily permeate wild-type NR1/NR2A channels, presumably because the head group of N1-OsSpm can pass through the narrowest part of the channel pore. N-Sulfonyl-polyamines such as N1-DnsSpm and N1-OsSpm represent a new class of polyamine antagonists with which to study glutamate receptor ion channels.

Amino Acid Sequence↗
PubMed1997

Effect of cyclosporin A on the expression of tissue kallikrein, kininogen, and bradykinin receptor in rat.

The tissue kallikrein-kinin system is involved in vasodilation and blood pressure regulation. In the present study, we investigated the effects of chronic cyclosporin A (CsA) administration on blood pressure and the expression of tissue kallikrein, kininogen, and bradykinin receptor in normotensive Wistar rats. Chronic administration of CsA significantly increased systolic blood pressure compared with control rats (n = 6, P < 0.01), although body weight was significantly lower than control rats (n = 6, P < 0.01). The development of hypertension was accompanied by the altered expression of kallikrein-kinin system components. Immunoreactive renal kallikrein and urinary excretion of tissue kallikrein levels were increased by chronic administration of CsA (n = 5 or 6, P < 0.05). Levels of N-tosyl-L-phenylalanine chloromethyl ketone-trypsin and kallikrein-releasable kininogens in sera increased in response to chronic CsA treatment (n = 5 or 6, P < 0.05). Chronic CsA treatment significantly increased renal kallikrein, bradykinin B2 receptor, and hepatic kininogen mRNA levels. The increased levels of tissue kallikrein-kinin system components were accompanied by significant increases in 24-h urine excretion and water intake after chronic CsA treatment (n = 5, P < 0.05). These results suggest that enhanced activity of the tissue kallikrein-kinin system may compensate for the CsA-induced vasoconstriction and hypertension.

Animals↗
PubMed1997

Kallikrein-kinin system and blood pressure sensitivity to salt.

We evaluated the blood pressure response to chronic salt loading in a rat strain inbred for low urinary kallikrein excretion. Low-kallikrein rats showed greater systolic blood pressure values (130 +/- 1 versus 114 +/- 2 mm Hg in controls; P < .05) at 9 weeks of age. Systolic blood pressure was increased after 10 days of dietary sodium loading in the low-kallikrein group and remained unchanged in controls (153 +/- 1 versus 112 +/- 2 mm Hg, P < .01). In additional experiments, blood pressure sensitivity to salt was tested in low-kallikrein rats receiving a chronic infusion of rat glandular kallikrein (1.7 micrograms/day per 100 g body weight, IV) or vehicle. Systolic blood pressure of vehicle-treated rats was increased by salt loading (from 138 +/- 1 to 158 +/- 2, 153 +/- 1, and 145 +/- 2 mm Hg at 5, 10, and 15 days, respectively; P < .01), while it remained unchanged in the kallikrein-treated group (from 136 +/- 2 to 146 +/- 5, 140 +/- 2, and 134 +/- 4 mm Hg at 5, 10, and 15 days, respectively; P = NS). Urinary kallikrein excretion was increased by kallikrein infusion (from 13.6 +/- 1.4 to 17.8 +/- 2.1 nanokatals per 24 hours; P < .01). Plasma immunoreactive kallikrein levels were higher in the kallikrein-treated group (66.4 +/- 4.4 versus 57.7 +/- 1.4 ng/mL in vehicle-treated rats; P < .05). On normal sodium diet, the ratio of kidney weight to body weight was lower in low-kallikrein rats (329 +/- 5 versus 370 +/- 8 mg/100 g body weight in controls; P < .01). This difference was associated with a decreased number of glomeruli per unit square area and increased width of Bowman's space. These results indicate that kallikrein replacement prevents the exaggerated blood pressure increase observed in rats with a genetically determined defect in urinary kallikrein excretion. Histological abnormalities are present at different levels in the nephron, and they may be functionally related to the altered cardiovascular and renal phenotype of this strain.

Animals↗
PubMed1997

Hypotension in transgenic mice overexpressing human bradykinin B2 receptor.

Bradykinin binds to its receptor at target organs and exerts a wide spectrum of biological activities including vasodilation, smooth muscle contraction and relaxation, pain, and inflammation. To gain a better insight into the physiological function of this potent vasoactive peptide, we created transgenic mice that harbor the human bradykinin B2 receptor transgene under the control of the Rous sarcoma virus 3'-LTR promoter (RSV-cHBKR). Expression of HBKR in these transgenic mice was identified in the aorta, brain, heart, lung, liver, kidney, uterus, and prostate gland by reverse transcription-polymerase chain reaction Southern blot analysis. Two transgenic mouse lines expressing the human B2 receptor resulted in a significant reduction of blood pressure (84.2 +/- 0.6 mm Hg, n = 28; 76.9 +/- 0.8 mm Hg, n = 24; P < .001) compared with the control littermates (96.9 +/- 0.4 mm Hg, n = 52). Administration of Hoe 140, a bradykinin B2 receptor antagonist, restored the blood pressure of the transgenic mice to normal levels within 1 hour, and the effect diminished within 4 hours. The transgenic mice displayed enhanced blood pressure-lowering effect induced by a bolus intra-aortic injection of kinin and showed increased response in kinin-induced uterine smooth muscle contractility compared with control littermates. These studies show that overexpression of human bradykinin B2 receptor causes a sustained reduction of blood pressure in transgenic mice. They also suggest that the B2 receptor-mediated signal transduction pathway plays a role in blood pressure regulation.

Adrenergic beta-Antagonists↗
PubMed1997

Prolonged reduction of high blood pressure with human nitric oxide synthase gene delivery.

Endothelium-derived nitric oxide (NO) in peripheral vessels has been shown to modulate vascular resistance and blood pressure. We explored the effect of a continuous supply of human endothelial NO synthase (eNOS) on the blood pressure of spontaneously hypertensive rats (SHR) by somatic gene delivery. A DNA construct containing the human eNOS gene fused to the cytomegalovirus promoter/enhancer was injected into SHR through the tail vein. A single injection of the naked eNOS plasmid DNA caused a significant reduction of systemic blood pressure for 5 to 6 weeks in SHR, and the effect continued for up to 10 to 12 weeks after a second injection. The differences were significant from 2 to 12 weeks postinjections (n=6, P<.01). In a separate experiment, L-arginine, the substrate of eNOS, was supplied in drinking water at a concentration of 7.5 g/L for 11 weeks after eNOS gene delivery. A maximal blood pressure reduction of 21 mm Hg in SHR was observed with eNOS DNA compared with that of control SHR injected with vector DNA (181.9+/-1.46 versus 202.7+/-2.79 mm Hg, mean+/-SEM, n=6, P<.01). Human eNOS gene delivery induces significant increases in urinary and aortic cGMP levels and urinary and serum nitrite/nitrate content (P<.05), while no significant differences in body weight, heart rate, water intake, food consumption, or urine excretion were observed. These results indicate that somatic delivery of the human eNOS gene induces a prolonged reduction of high blood pressure and raises the potential of using eNOS gene therapy for hypertension and cardiovascular diseases.

Animals↗
PubMed1997

Signal amplification in the detection of single-copy DNA and RNA by enzyme-catalyzed deposition (CARD) of the novel fluorescent reporter substrate Cy3.29-tyramide.

We demonstrate that the CAtalyzed Reporter Deposition method (CARD), utilizing the novel fluorescent reporter Cy3.29-tyramide, is successful in the Fluorescent in Situ Hybridization (FISH) detection of RNA and single-copy DNA. Histone 4 expression is detected in RNA extracts of 5-phase, synchronized HeLa cells by dot-blot analysis. Gene expression of histone 4 in HeLa cells is demonstrated by FISH via CARD, utilizing oligonucleotide probes. Fluorescence intensity measurements on CARD-amplified histone 4 RNA detection showed (a) a 25-fold amplification of the signal brightness by biotinylated oligonucleotide probes and (b) a sixfold amplification of the signal brightness by horseradish peroxidase (HRP)-labeled histone 4 probes vs the directly stained control. The sensitivity of the CARD method is demonstrated by the FISH detection of single-copy DNA on human corneal fibroblast and HeLa S5 interphase nuclei. Chromosomal localization of the single copy DNA is demonstrated on HeLa S3 metaphase chromosome spreads.

Avidin↗
PubMed1997

Adrenomedullin gene delivery reduces blood pressure in spontaneously hypertensive rats.

Adrenomedullin (ADM) is a potent vasoactive peptide. In this study, we explored the effects of a continuous supply of ADM by somatic gene delivery on spontaneously hypertensive rats (SHR). DNA constructs containing the human ADM cDNA fused to either the cytomegalovirus promoter (CMV-cADM) or Rous Sarcoma virus 3'-long terminal repeat (RSV-cADM) were intravenously injected into SHR through the tail vein. Expression of human ADM in SHR was identified in the kidney, adrenal gland, heart, and lung by radioimmunoassay and reverse transcription-polymerase chain reaction followed by Southern blot analysis. A single injection of ADM plasmid DNA in young adult SHR (7 wk old) caused a significant reduction in systolic blood pressure for up to 5 wk (p < 0.05). A second injection of CMV-cADM 5 wk after the first delivery resulted in a further reduction in blood pressure for another 3 wk (p < 0.001). A maximal blood pressure reduction of 22 mmHg in SHR was observed 7 wk after injection of CMV-cADM plasmid DNA (185 +/- 1.7 mmHg, n = 6, p < 0.001), and a reduction of 15 mmHg was observed after injection of RSV-cADM (192 +/- 2.7 mmHg, n = 6, p < 0.001), as compared with control rats given vector DNA (207 +/- 2.4 mmHg, n = 6). Similarly, injection of CMV-cADM plasmid DNA in adult SHR (10 wk old) resulted in a significant reduction in blood pressure for up to 6 wk. Antibodies to either human ADM or its plasmid DNA were not detected in rat sera after the second injection. These studies indicate that intravenous injection of the human ADM gene in hypertensive rats results in expression of the foreign gene and induces a long-lasting reduction in blood pressure.

Adrenomedullin↗
PubMed1997

Kallikrein-binding protein levels are reduced in the retinas of streptozotocin-induced diabetic rats.

PURPOSE: To determine the involvement of rat kallikrein-binding protein (RKBP) in the development of diabetic retinopathy. METHODS: Diabetes was induced by streptozotocin (STZ) (55 mg/kg body weight in 0.05 M citrate buffer, pH 4.5) in male Sprague-Dawley rats (150 to 175 g, 6 weeks old) as confirmed by hyperglycemia and reduced body weight. Retinas were dissected from animals at 1, 2, and 4 months of diabetes. The functional activity of RKBP in retinal homogenates was determined by its complex formation with tissue kallikrein. Immunoreactive RKBP levels were measured by enzyme-linked immunosorbent assay. The RKBP messenger RNA (mRNA) levels in the retina were measured by Northern blot analysis using the RKBP complementary DNA probe. The activity of total Na+,K(+)-ATPase was determined by a radioassay. Total protein concentration was determined by a protein assay. RESULTS: The kallikrein-binding activity was reduced in the retinas of STZ-diabetic rats at 1 (59%), 2 (50%), and 4 (38%) months of diabetes compared to those of age-matched control subjects. Levels of immunoreactive RKBP were significantly lower in the diabetic animals at each time point examined compared to those of control subjects. At 1 and 2 months of diabetes, RKBP levels (nanogram/milligram protein) were decreased significantly to 6.9 +/- 0.7 (n = 8) and 10.6 +/- 1.0 (n = 8), respectively, compared to those of age-matched control subjects (14.1 +/- 0.7, n = 8, P < 0.001, and 14.1 +/- 1.2, n = 8, P < 0.01). At 4 months of diabetes, retinal RKBP levels were lower in both control and diabetic groups, but RKBP levels in diabetic groups were significantly lower (5.8 +/- 0.6, n = 8) than those of the age-matched control subjects (8.4 +/- 0.9, n = 8, P < 0.01). Similarly, Northern blot analysis showed that RKBP mRNA levels were reduced in the retina of each group of STZ-diabetic rats, suggesting that the decrease in RKBP occurred at the level of transcription. CONCLUSIONS: The results show that STZ-induced diabetic rats have decreased retinal RKBP; moreover, this suggests that RKBP may contribute to diabetic retinopathy.

Animals↗
PubMed1997

Kallikrein gene therapy in newborn and adult hypertensive rats.

The tissue kallikrein-kinin system has been postulated to play an important role in blood pressure regulation. Kallikreins are serine proteinases that release potent vasodilating kinin peptides from precursor kininogens by limited proteolysis. Our recent studies show that systemic delivery of the human tissue kallikrein gene into adult spontaneously hypertensive rats (SHR) results in a sustained reduction of blood pressure for several weeks. The goal of this study is to evaluate whether early delivery of the kallikrein gene into newborn SHR could exert a suppressive effect on blood pressure phenotype during rat growth and development. A human tissue kallikrein cDNA construct, under the control of cytomegalovirus promoter (CMV-cHK), or vector DNA was injected subcutaneously into the necks of 2-day-old SHR. Blood pressures were monitored biweekly from 3 to 19 weeks by the tail-cuff method. A single injection of the human kallikrein cDNA construct caused a significant reduction of blood pressure (n = 6, p < 0.001) from 11 to 17 weeks after injection compared with control rats receiving vector DNA. Intravenous delivery of the human tissue kallikrein gene into adult SHR produced blood pressure lowering effects (n = 6, p < 0.001) that lasted for 6 weeks in male but not in female rats. The expression of human tissue kallikrein in rats was identified by reverse transcription polymerase chain reaction followed by Southern blot analysis and an ELISA specific for human tissue kallikrein. Kallikrein gene delivery did not cause any changes in body weight, urine volume, or water intake in the experimental animals compared with the control group. No antibodies to either human tissue kallikrein or its DNA were detected in rat sera 19 weeks postinjection. These results show that delivery of the kallikrein gene at an early stage of life has a protective effect against development of hypertension in adult SHR and that gender differences could be a factor in kallikrein gene therapy for the treatment of hypertensive disorders.

Aging↗
PubMed1997

The enzymic potential of tissue kallikrein (rK1) in rat submandibular saliva depends on whether it was secreted via constitutive or regulated pathways.

The enzymic activity and immunoreactivity of rat tissue kallikrein (rK1) secreted at rest by granular duct cells of unstimulated submandibular glands has been compared with that secreted on autonomic nerve stimulation. Although a direct vesicular, constitutive secretory pathway operates for rK1 secretion from granular duct cells of unstimulated and parasympathetically stimulated glands the rK1 was not present in a pro-form and actually showed a greater enzymic activity per unit immunoreactive protein than the granule-derived rK1 in sympathetically evoked saliva. Constitutively secreted rK1 was found to be in a single chain molecular form by reducing SDS gel electrophoresis. In contrast rK1 secreted from the storage granule pool of granular duct cells on sympathetic nerve stimulation was present in much higher amounts and occurred in both one-chain and two-chain forms as revealed by SDS gel electrophoresis under reducing conditions. The lower enzymic potential of rK1 in sympathetically evoked saliva might be accounted for by its conversion to a two-chain form.

Animals↗
PubMed1996

Tissue kallikrein-binding protein reduces blood pressure in transgenic mice.

The kallikrein-kinin system participates in blood pressure regulation. One of the kallikrein-kinin system components, kallikrein-binding protein, binds to tissue kallikrein and inhibits its activity in vitro. To investigate potential roles of rat kallikrein-binding protein (RKBP) in vivo, we have developed transgenic mice that express an RKBP gene under the control of the mouse metallothionein metal-responsive promoter. Expression of the transgene, RKBP, was detected in the liver, kidney, lung, heart, pancreas, salivary glands, spleen, brain, testis, and adrenal gland at the mRNA and protein levels. Systolic blood pressures of homozygous transgenic mice were 88.5 +/- 0.8 mm Hg (mean +/- S.E., n = 19, P < 0.001) for one line and 88.8 +/- 1.6 mm Hg (mean +/- S.E., n = 19, P < 0.001) for another, as compared with 100.5 +/- 0.8 mm Hg (mean +/- S.E., n = 18) for control mice. Direct blood pressure measurements of these transgenic mice through an arterial cannula showed similar reductions of blood pressure. Intravenous injection of purified RKBP into mice via a catheter produced a dose-dependent reduction of the mean arterial blood pressure. Our findings suggest that RKBP may function as a vasodilator in vivo, independent of regulating the activity of tissue kallikrein.

Animals↗