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Biomedical subjects

R C Harris

Publications and source records attributed to R C Harris.

At least 91 records · Page 5Linked to original sources
PubMed1995

Physiological, metabolic and biochemical responses of horses competing in the speed and endurance phase of a CCI*****3-day-event.

The present study was undertaken to investigate physiological, metabolic, haematological and biochemical changes in horses competing in the Speed and Endurance test of a Concours Complet International (CCI)*****3-day-event held under FEI rules. A total of 28 horses competing in the Burghley Horse Trials Speed and Endurance test were selected to be monitored: 11 horses in 1993 and 17 horses in 1994. Of the 28 horses selected, 17 completed the Speed and Endurance test and went on to complete the showjumping test. Mean +/- s.d. shade temperature and relative humidity, black globe temperature and wind speed were 13 +/- 1 and 20 +/- 2 degrees C, 54 +/- 3 and 55 +/- 10%, 17 +/- 2 and 29 +/- 4 degrees C and 2.7 +/- 0.7 and 1.2 +/- 0.3 m/s, for 1993 and 1994, respectively. Mean heart rate during Phases A, B and D was not significantly different between years, but mean heart rate during Phase C and X was significantly higher in 1994. Mean (+/- s.d.) heart rate on Phase B and D for all horses in both 1993 and 1994 was 198 +/- 8 and 188 +/- 11 beats/min, respectively. Mean heart rate during Phase D showed a poor correlation with mean speed (r = 0.412). Total mean (+/- s.d.) weight loss from the start of Phase A to the end of Phase D was 15.5 +/- 6.1 kg in 1993 and 16.5 +/- 5 kg in 1994 and did not differ significantly between years. Following 14-18 h completion of Phase D, mean bodyweight was not significantly different from that at the start of Phase A in either year. Mean rectal temperature at the end of Phase D was 41 +/- 0.6 degrees C and 41.1 +/- 0.6 degrees C in 1993 and 1994, respectively (P > 0.05). Both the lowest (39.7 degrees C) and highest (41.8 degrees C) rectal temperatures were recorded at the end of Phase D in 1994. Plasma lactate concentrations at the end of Phase D were 8.5-38.5 mmol/l. The highest lactate concentration also coincided with the highest plasma glucose concentration (11.4 mmol/l) as well as the joint fastest time in either year, although overall lactate showed only weak correlations with mean speed on Phase D (r = 0.12, 1993; r = 0.58, 1994). While the Speed and Endurance test at CCI*****level run in a temperate climate presents a considerable challenge to the fitness and ability of the horses competing, the metabolic and physiological changes are not extreme. The majority of horses that finish the test appear to undergo a rapid and considerable degree of recovery and are able to present sound at the final inspection, take part in the showjumping test and complete the competition.

Animals↗
PubMed1995

Signaling and growth responses of LLC-PK1/Cl4 cells transfected with the rabbit AT1 ANG II receptor.

Angiotensin II (ANG II) receptors of the AT1 subtype are present on the apical and basolateral membranes of renal proximal tubule cells. Cells of the proximal tubulelike cell line, LLC-PK1/Cl4, were transfected with an expression plasmid containing cDNA encoding the rabbit AT1 ANG II receptor. In transfected cells, specific binding of 125I-ANG II was detected on both apical and basolateral membranes; wild-type LLC-PK1/Cl4 cells did not express ANG II receptors. In transfected cells, apical or basolateral ANG II increased both S6 kinase activity and incorporation of [3H]leucine. In cells pretreated with pertussis toxin, the stimulatory effect of apical or basolateral ANG II on [3H]leucine incorporation was abolished. In contrast, ANG II did not affect mitogenesis, determined by [3H]thymidine incorporation. Apical or basolateral ANG II (10(-6) M) stimulated phosphoinositide turnover by 13.4 +/- 4.4% (n = 8) and 16.3 +/- 4.2% (n = 9), respectively. The activity of protein kinase C, determined by phosphorylation of a specific protein kinase C peptide substrate, was also stimulated by ANG II in transfected cells. Apical or basolateral ANG II had no significant effect on cellular adenosine 3',5'-cyclic monophosphate levels. In permeabilized transfected cells, apical ANG II (10(-6) M) inhibited the phosphorylation of a specific peptide substrate of protein kinase A; lower apical concentrations or basolateral ANG II were without significant effect. These results indicate that AT1 ANG II receptors sort to both apical and basolateral membranes in renal epithelial cells and are coupled to activation of phospholipase C. ANG II stimulates protein synthesis by binding to either apical or basolateral receptors; this effect requires coupling to G proteins and may be mediated by activation of S6 kinase. Because high concentrations of ANG II exist in proximal tubule, binding to apical and basolateral receptors may regulate proximal tubule cell growth under physiological conditions.

Animals↗
PubMed1995

Polarized rabbit type 1 angiotensin II receptors manifest differential rates of endocytosis and recycling.

Receptor-mediated endocytosis and recycling have been described for extrarenal angiotensin II (ANG II) receptors. In proximal tubule (PT) epithelia expressing polarized ANG II receptors, these processes have not been examined as thoroughly. We utilized a PT cell model, LLC-PKCl4 cells stably transfected with rabbit type 1 ANG II receptor (AT1R) cDNA, to investigate these properties. LLC-PK-AT1R cells expressed the rabbit AT1R transcript and displayed losartan-inhibitable specific 125I-labeled ANG II binding at apical (AP) and basolateral (BL) membranes when grown on permeable supports. AP AT1R internalized 125I-ANG II more rapidly than BL AT1R, and phenylarsine oxide treatment inhibited AP AT1R internalization without significantly affecting BL AT1R endocytosis. Pertussis toxin had no effect on AP or BL AT1R endocytosis. In addition, AP AT1R recovered specific 125I-ANG II binding after ANG II treatment (a measure of recycling). BL AT1R displayed minimal recovery of 125I-ANG II binding after ANG II pretreatment. These data suggested that AP AT1R enter endocytic/endosomal pathways. Phospholipase A2 (PLA2) activity has been linked to endosomal fusion in other systems, and PT brush-border membrane AT1R also have been associated with PLA2 activity. LLC-PK-AT1R cells were therefore treated with quinacrine, a nonspecific PLA2 inhibitor, or Compound I (CI), a selective Ca(2+)-independent PLA2 inhibitor, to determine if PLA2 activity was linked to AT1R recycling. Both quinacrine and CI decreased AP AT1R recycling without affecting BL AT1R recycling. Polarized AT1R in LLC-PKCl4 cells thus display differential rates of endocytosis and recycling.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Role of cytochrome P-450 epoxygenase metabolites in EGF signaling in renal proximal tubule.

Epidermal growth factor (EGF) is a potent epithelial cell mitogen and induces eicosanoid production in many cell types. The present study examined signaling mechanisms for EGF in the renal proximal tubule, where high concentrations of cytochrome P-450 epoxygenase have been reported. In primary cultures of rabbit proximal tubule cells, EGF (30 nM) increased endogenous epoxyeicosatrienoic acid (EET) levels 5.3 +/- 1.4-fold within 10 min (n = 6). In these cells EGF-stimulated [3H]thymidine incorporation was significantly inhibited by the cytochrome P-450 inhibitors ketoconazole or clotrimazole but not by the cyclooxygenase inhibitor indomethacin. In fura 2-loaded proximal tubule cells, EGF caused a concentration-dependent increase in cytosolic Ca2+ concentration ([Ca2+]i), due to Ca2+ influx, which was inhibited by either ketoconazole or SKF-525A but not by indomethacin. Addition of 5,6-EET (0.5 microM) also induced Ca2+ influx in proximal tubule cells, whereas 8,9-11,12-, or 14,15-EET did not. In cells treated with bis(2-amino-5-methylphenoxy)ethane N,N,N',N'-tetraacetic acid tetraacetoxy-methyl ester to chelate [Ca2+]i, EGF-stimulated [3H]thymidine incorporation. These results indicate that EGF increases EET levels in proximal tubule and suggest that 5,6-EET or its metabolites may be a modulator of EGF-induced [Ca2+]i increases and involved in mitogenesis.

8,11,14-Eicosatrienoic Acid↗
PubMed1995

Angiotensin II upregulates type-1 angiotensin II receptors in renal proximal tubule.

Angiotensin II (Ang II) is an important regulator of proximal tubule salt and water reabsorption. Recent studies indicate that rabbit proximal tubule angiotensin II receptors are the type-1 (AT1R) subtype. We studied the effect of Ang II on proximal tubule receptor expression. Rabbits were treated with either angiotensin converting enzyme inhibitors or a low salt diet to modulate endogenous Ang II levels. In captopril-treated rabbits, liver and glomerular AT1R mRNA levels increased 242 +/- 125 and 141 +/- 60%, respectively (n = 6-7; P < 0.05), as determined by quantitative PCR. In contrast, proximal tubule AT1R mRNA levels decreased 40 +/- 11% (n = 6; P < 0.05). Binding of 125I Ang II to renal cortical basolateral membranes of captopril-treated rabbits decreased from 2.9 +/- 0.55 to 1.4 +/- 0.17 fmol/mg protein (n = 6; P < 0.025). In rabbits fed a sodium chloride-deficient diet for 4 wk, AT1R mRNA levels decreased 52 +/- 11% in liver and 43 +/- 7% in glomeruli (n = 4-5; P < 0.05), whereas they increased 141 +/- 85% (n = 5; P < 0.05) in proximal tubule. In basolateral membranes from rabbits on the sodium chloride-deficient diet, specific binding of 125I Ang II increased from 2.1 +/- 0.2 to 4.3 +/- 1.1 fmol/mg protein (n = 7; P < 0.05). To determine whether Ang II directly regulates expression of proximal tubule AT1 receptors, further studies were performed in cultured proximal tubule cells grown from microdissected S1 segments of rabbit proximal tubules and immortalized by transfection with a replication-defective SV40 vector. Incubation of these cells with Ang II (10(-11) to 10(-7) M) led to concentration-dependent increases in both AT1R mRNA levels and specific 125I Ang II binding. Pretreatment with pertussis toxin inhibited Ang II stimulation of AT1R mRNA. AT1R mRNA expression was decreased by either forskolin or a nonhydrolyzable cAMP analogue (dibutryl cAMP). Simultaneous Ang II administration overcame the inhibitory effect of forskolin but not dibutryl cAMP. These results indicate that proximal tubule AT1R expression is regulated by ambient Ang II levels, and Ang II increases AT1R mRNA at least in part by decreasing proximal tubule cAMP generation through a pertussis toxin-sensitive mechanism. Upregulation of proximal tubule AT1R by Ang II may be important in mediating enhanced proximal tubule sodium reabsorption in states of elevated systemic or intrarenal Ang II.

Angiotensin II↗
PubMed1995

Induction of heparin-binding epidermal growth factor-like growth factor mRNA in rat kidney after acute injury.

Previous studies have suggested that EGF or other members of the EGF family of mitogenic proteins are involved in proliferation of renal tubular epithelial cells occurring during recovery from injury to the kidney. The present studies examined whether expression of mRNA for the recently identified heparin-binding EGF-like growth factor (HB-EGF) is regulated in response to renal injury induced by either ischemia/reperfusion or mercuric chloride. Increased expression of HB-EGF mRNA was demonstrated in the post-ischemic kidney within 45 min of unilateral ischemia/reperfusion in the rat. Induction of HB-EGF mRNA occurred only when ischemia was followed by reperfusion, and was not eliminated by removal of blood cells from the post-ischemic kidney by saline perfusion. In situ hybridization with 35S-labeled antisense riboprobes of HB-EGF indicated that compared with control, there was increased HB-EGF mRNA expression in the 6 h post-ischemic kidney in the inner cortex and outer medulla in a patchy distribution, with the greatest expression in the inner stripe of the outer medulla. Expression occurred primarily in tubular epithelial cells. Recombinant human HB-EGF stimulated [3H]-thymidine incorporation in both primary cultures of rabbit proximal tubule cells and NRK 52E normal rat kidney epithelial cells, with potency similar to that of EGF. Induction of HB-EGF mRNA was observed in tubules freshly isolated from rat renal cortex or outer medulla when the tubules were subjected to reoxygenation after incubation in anoxic conditions. The nephrotoxin, mercuric chloride, also caused induction of HB-EGF mRNA both in vivo and in isolated rat cortical tubules. The anoxia/reoxygenation-induced expression of HB-EGF mRNA in isolated tubules was inhibited by the free radical scavengers, di- and tetra-methylthiourea, indicating involvement of reactive oxygen species. These findings indicate that HB-EGF mRNA is inducible in the kidney in vivo by acute tubular injury and suggest that HB-EGF may act as an autocrine/paracrine growth factor involved in proliferation of tubular epithelial cells and repair of the kidney.

Animals↗
PubMed1995

Angiotensin II-mediated renal injury.

During the past decade, experimental and clinical evidence has indicated an important role for the renin-angiotensin system in the progressive destruction of nephrons in a wide variety of chronic renal diseases. Studies have indicated that in the subtotally nephrectomized rat model of progressive glomerulosclerosis, in experimental diabetes mellitus, in the chronic phase of puromycin aminonucleoside-induced nephrotic syndrome and in Heymann's nephritis, angiotensin-converting enzyme (ACE) inhibitors dramatically preserve both nephron structure and function. Clinical studies have similarly noted that chronic administration of ACE inhibitors inhibits progression of renal failure in type I diabetes and type II diabetes as well as primary glomerulopathies, sickle cell nephropathy, systemic lupus erythematosis, chronic pyelonephritis and adult polycystic kidney disease. Current evidence suggests that the beneficial effect of ACE inhibitors is primarily due to inhibition of angiotensin II production, and there is strong suggestive evidence for increases in local intrarenal activation of the renin-angiotensin system in these conditions. In obstructive uropathy, activation of the renin-angiotensin system has also been shown to be an important aspect of the early functional changes and may be of importance in the subsequent generation of interstitial fibrosis. In the obstructed kidney, renin and angiotensinogen production increase and type I angiotensin receptors decrease. Inhibitors of angiotensin II production and angiotensin II action partially reverse the vasoconstriction and the reduced renal blood flow, and abolish the changes in expression of AT1 MRNA induced by obstruction. Studies suggest that the angiotensin-mediated increases in tubulointerstitial fibrosis may be mediated by increased production of transforming growth factor-beta.

Acute Kidney Injury↗
PubMed1995

Clinical observations made in nonheat acclimated horses performing treadmill exercise in cool (20 degrees C/40%RH), hot, dry (30 degrees C/40%RH) or hot, humid (30 degrees C/80%RH) conditions.

Four horses (H, J, N and M) undertook a treadmill competition exercise test (CET), designed to simulate the physiological and metabolic stresses of the Speed and Endurance phase of a 3-day-event, under 3 different environmental conditions: 20 degrees C/40% relative humidity (RH) (cool, dry [CD]: 2 sessions); 30 degrees C/40%RH (hot, dry [HD]) and 30 degrees C/80%RH (hot, humid [HH]) (Marlin et al. 1995). A number of subjective clinical observations were made at designated time points throughout the exercise test and initial recovery period including buccal mucous membrane colouration, capillary refill time, neck and point of shoulder skin pinch recovery time, grade of abdominal sounds; anal sphincter tone as well as the presence or absence of fatigue and ataxia. The aim was to investigate their value in predicting performance in the final canter phase of the CET equivalent to the cross-country or Phase D of a field competition. In addition, the use of a more objective assessment, the cardiac recovery index (CRI), was investigated together with the heart rate, rectal temperature and respiratory frequency at the end of Phase C and at the 8 min point of the 10 Minute Box (8'X). The CRI was calculated according to the formula CRI = P2-P1 where P2 = the heart rate in beats/min at the 8 min point of the '10 Minute Box' (Phase X) of the CET. P1 = the heart rate (beats/min) at the 7 min point just before the horse was made to trot over a distance of 80 m at a speed of 3.7 m/s (at a 3 degrees incline) before returning to a walk. The study suggested that the subjective tests carried out at the 'End-C' and/or '8'X' time points were not useful in predicting subsequent performance in the final canter phase (Phase D) and neither were heart rate, rectal temperature or respiratory frequency. However, the only horse (Horse H) to complete the full CET under HH conditions was the only animal to show a decrease in respiratory frequency between the End-C and 8'X time points. All others showed an obvious increase. Under HH conditions, Horse H also had the lowest CRI. For 3 of the horses the highest CRI value was found under the HH conditions, for the fourth horse an equally high CRI value was found with one of the CD sessions. However, under the HH conditions, both P1 and P2 values were > 100. The study suggested that it could be beneficial if a suitably modified CRI test, as well as a procedure to monitor the change in respiratory frequency during the 10 Minute Box, were evaluated further on the treadmill and in the field with respect to their potential usefulness as additional aids to the assessment of a horse's suitability to proceed to Phase D.

Acclimatization↗
PubMed1995

Electrolyte and total protein changes in nonheat acclimated horses performing treadmill exercise in cool (20 degrees C/40%RH), hot, dry (30 degrees C/40%RH) or hot, humid (30 degrees C/80%RH) conditions.

Four horses (H, J, N and M) undertook a simulated competition exercise test (CET), designed to simulate physiological and metabolic stresses of the Speed and Endurance (S & E) test of a 3-day-event, under 3 different environmental conditions: 20 degrees C/40% relative humidity (RH) (cool, dry [CD] 2 sessions); 30 degrees C/40% RH (hot, dry [HD]) and 30 degrees C/80% RH (hot, humid [HH]) (Marlin et al. 1995a). Venous blood samples for electrolyte and total protein (TP) determinations were collected from indwelling catheters at predetermined time points throughout each CET and initial 30 min recovery period. Venous blood samples were collected by jugular venepuncture at 2 h and 24 h after the end of the final 8 min canter (Phase D). The effects of exercise, environmental condition and horse on venous TP, sodium (Na+), potassium (K+), chloride (Cl-), calcium (Ca2+), magnesium (Mg2+) and phosphate (PO4(3-)) plasma concentrations were investigated. In addition, the effect of environmental condition on estimated cation loss was evaluated. All horses completed the full CET under the CD and HD conditions, but only one horse completed the full 8 min of the final canter Phase D under HH conditions. Exercise had a significant (P < 0.05) effect on all parameters similar to those reported previously in field competitions. There was a significant (P < 0.05) interaction between time and horse for TP, Na+ and Cl-. Overall, the environmental condition had a significant (P < 0.05) effect on all parameters, but the differences were not considered to be of any physiological relevance, other than for Cl- and TP. There was a significant (P < 0.05) interaction between time and environmental condition for Cl- and TP only. During much of the CET and recovery period, mean Cl- values were higher with the first CD session than the second CD session or under the HH or HD conditions. For TP after 2 min of Phase D and during the initial recovery period, concentrations were higher under the HH conditions and returned to the Pre- values less quickly. There were marked individual variations in the estimated cation losses and no consistent effect of environmental condition was found. Although estimated fluid loss was similar following both HD and HH sessions, restoration of bodyweight was slower following the CET studies under HH conditions.

Acclimatization↗
PubMed1994

Skeletal muscle characteristics in 2 year-old race-trained thoroughbred horses.

Skeletal muscle samples were obtained by needle biopsy from two depths of the m. gluteus medius of 50, young race-trained thoroughbred racehorses. Histochemical and biochemical characteristics of the muscle samples were analysed. Fibres were classified as type I, type IIa or type IIb on the basis of the pH dependent lability of the myosin ATPase reaction. The activities of citrate synthase (CS) and glycogen phosphorylase (Phos) were determined. Muscle fibre composition varied markedly between deep and superficial muscle samples and this was reflected in differences in the activities of citrate synthase (CS) and phosphorylase (Phos). CS activity was greater in samples taken from a depth of 90 mm (deep) than those taken from a depth of 40 mm (superficial: 122 +/- 19 compared with 88 +/- 16 mumol/g dry muscle/min at 25 degrees C). Phos activity was greater in superficial samples (137 +/- 20) compared with deep samples (117 +/- 21). Regression analysis was used to estimate the enzyme activities in the different fibre types. No significant correlations were observed between histochemical and biochemical measures and subsequent racing performance.

Animals↗
PubMed1994

Extrarenal complications of the nephrotic syndrome.

The systemic complications of nephrotic syndrome are responsible for much of the morbidity and mortality seen with this condition. This review discusses the causes for the hypoalbuminemia and the associated metabolic abnormalities of the nephrotic syndrome. No unifying hypothesis exists for the induction, maintenance, and resolution of nephrotic edema. In view of the wide spectrum of renal diseases leading to the nephrotic syndrome, more than a single mechanism may be responsible for the renal salt retention in these diverse conditions. Although hypoalbuminemia may be important, especially when plasma oncotic pressure is very low (serum albumin < 1.5 to 2.0 g/dL), primary impairment of salt and water excretion by the nephrotic kidney appears to be a major factor in pathogenesis of the edema. However, the decreased serum albumin and/or oncotic pressure seen with nephrotic syndrome is a major contributing factor to the development of the hyperlipidemia of nephrotic syndrome. Patients with unremitting nephrotic syndrome should be considered for combined dietary and lipid-lowering drug therapy. Urinary losses of binding proteins lead to the observed abnormalities in the endocrine system and in trace metals, and urinary losses of coagulation factors contribute to the hypercoagulable state. At present, selective renal venography is recommended when the suspicion of renal vein thrombosis is justified by clinical presentation. The impact on renal function caused by treating asymptomatic chronic renal vein thrombosis is undetermined, but anticoagulation for chronic renal vein thrombosis is associated with relatively few complications.

Acute Kidney Injury↗
PubMed1994

Reduced proximal tubule angiotensin II receptor expression in streptozotocin-induced diabetes mellitus.

Diabetes mellitus is characterized by alterations in the intrarenal renin-angiotensin system, including decreases in glomerular angiotensin II (Ang II) receptor density. Since Ang II regulates proximal tubule transport function, the present studies examined whether diabetes altered expression of proximal tubule receptors. In basolateral membranes from 14 day streptozotocin-induced diabetic rats, specific binding of 125I Ang II was decreased to 53 +/- 8% of control (3.2 +/- 0.5 vs. 1.5 +/- 0.2 fmol/mg protein; N = 7; P < 0.02). Similarly, in proximal tubule brush border membranes from diabetic animals, specific binding was decreased to 63 +/- 11% of control (1.1 +/- 0.2 vs 0.6 +/- 0.1 fmol/mg protein; N = 9; P < 0.05). Concomitant insulin treatment reversed the decrease in specific binding of 125I Ang II to basolateral membranes (109 +/- 26% of control; N = 3) and to brush border membranes (85 +/- 17% of control; N = 6). In order to determine if changes in expression of type-1 Ang II receptors (AT1R) accompanied the changes in binding, quantitative polymerase chain reaction of AT1R mRNA was performed and expressed as the ratio of the amplified AT1R to that of an Msc1/Msc1 internal deletion mutant and normalized to that of beta-actin. In total RNA from proximal tubule suspensions of diabetic animals, AT1R mRNA expression decreased by 38% (21 +/- 3 vs. 13 +/- 2 cpm AT1R/cpm deletion mutant/cpm beta actin/10(6); N = 4; P < 0.0025). Insulin treatment reverted AT1R mRNA expression to control levels (22 +/- 3 cpm AT1R/cpm deletion mutant/cpm beta actin/10(6); P < 0.001 compared to the untreated group).(ABSTRACT TRUNCATED AT 250 WORDS)

Actins↗
PubMed1994

Mechanical stretch/relaxation of cultured rat mesangial cells induces protooncogenes and cyclooxygenase.

In cultured rat glomerular mesangial cells, continuous cycles of stretching and relaxation (stretch/relaxation) stimulate cell proliferation, protein synthesis, and prostaglandin production. We examined regulation of gene expression that may underlie these alterations in cell functions. Stretch/relaxation caused time-dependent induction of the immediate early genes, c-fos and zif 268/egr-1, with maximal increases occurring between 15 and 30 min. The mitogen-inducible prostaglandin G2/H2 synthase (PGH2S-2) gene was also induced within 30 min of stretch/relaxation, with concomitant increases in the immunoreactive PGH2S-2 protein. These gene inductions were preceded by transient translocation of protein kinase C activity from cytosol to membrane as well as by increases in 45Ca2+ uptake and total cellular calcium content. The stretch/relaxation-induced expression was suppressed by protein kinase C inhibition, whereas less profound inhibition was observed with inhibition of calcium influx in low (100 nM) calcium buffer. These findings indicate that in mesangial cells mechanical stress induces expression of the protooncogenes and the mitogen-inducible cyclooxygenase primarily through protein kinase C-dependent mechanisms.

Animals↗
PubMed1994

Cyclooxygenase-2 is associated with the macula densa of rat kidney and increases with salt restriction.

The kidney is a rich source of prostaglandins. These eicosanoids, formed by cyclooxygenase-dependent metabolism of arachidonic acid, are important physiologic mediators of renal glomerular hemodynamics and tubular sodium and water reabsorption. Two separate isoforms of cyclooxygenase (COX) have now been identified: constitutive COX-1, encoded by a 2.8-kb mRNA, and mitogen-activated COX-2, encoded by a 4.0-4.5-kb mRNA. COX-2 expression increases during development and inflammation, but, except for brain, constitutive expression is low. It has been generally accepted that physiologic renal production of prostaglandins is mediated by COX-1. However, in the absence of inflammation, low levels of COX-2 mRNA are also detectable in the kidney. To examine the role of COX-2 in the kidney and determine its intrarenal localization, we used a 1.3-kb cDNA probe specific for the 3' untranslated region of rat COX-2 and COX-2-specific antiserum. The COX-2-specific cDNA probe hybridized with a 4.4-kb transcript in total RNA from adult rat kidney. Immunoblots of microsomes isolated from kidney cortex and papilla indicated immunoreactive COX-2 in both locations. In situ hybridization and immunohistochemistry indicated that renal cortical COX-2 expression was localized to the macula densa of the juxtaglomerular apparatus and to adjacent epithelial cells of the cortical thick ascending limb of Henle. In addition, COX-2 immunoreactivity was detected in interstitial cells in the papilla. No COX-2 message or immunoreactive protein was detected in arterioles, glomeruli, or cortical or medullary collecting ducts. When animals were chronically sodium restricted, the level of COX-2 in the region of the macula densa increased threefold (from 0.86 +/- 0.08 to 2.52 +/- 0.43/mm2) and the total area of the COX-2 immunoreactive cells in cortex increased from 34 microns2/mm2 of cortex to 226 microns2/mm2 of cortex. The intrarenal distribution of COX-2 and its increased expression in response to sodium restriction suggest that in addition to its proposed role in inflammatory and growth responses, this enzyme may play an important role in the regulation of salt, volume, and blood pressure homeostasis.

Animals↗
PubMed1993

Cyclooxygenase-derived metabolites of 8,9-epoxyeicosatrienoic acid are potent mitogens for cultured rat glomerular mesangial cells.

The mitogenic effects of 11(R)-hydroxy-8,9-epoxyeicosatrienoic acid (EET) enantiomers were investigated in cultured rat glomerular mesangial cells. Both 11(R)-hydroxylated 8(R),9(S)- and 8(S),9(R)-EET at 1 microM stimulated [3H]-thymidine incorporation to 300% and 280%, with 50% maximal effect occurring at 8 x 10(-9) M and 1 x 10(-8) M, respectively. Similar concentration-dependent effects were observed in stimulating induction of the immediate early gene, c-fos. Mitogenic activity of the 11(R)-hydroxylated enantiomers was not affected by prior downregulation of protein kinase C, suggesting involvement of protein kinase C-independent mechanisms. These findings suggest that either trans- or intracellular metabolism of 8,9-EET by cyclooxygenase occurs during inflammatory glomerular diseases and that the resulting metabolites are involved in mesangial cell proliferation.

8,11,14-Eicosatrienoic Acid↗
PubMed1993

Hepatocyte growth factor stimulates phosphoinositide hydrolysis and mitogenesis in cultured renal epithelial cells.

Hepatocyte growth factor (HGF), a novel heparin-binding peptide growth factor of MW 97-kDa, is a potent mitogen for parenchymal hepatocytes. HGF is present in normal serum and increases following liver injury or partial hepatectomy. In addition to liver, HGF mRNA has been detected in kidney. In cultured rabbit proximal tubule cells, recombinant human HGF (10(-10) M) increased DNA synthesis, measured as [3H] thymidine incorporation, from 1345 +/- 213 to 2931 +/- 636 cpm/10(6) cells; n = 9; p < 0.005). HGF was found to exert mitogenic effects at lower concentrations than epidermal growth factor (EGF), with half maximal effects seen at 6 x 10(-11) M compared to 7 x 10(-10) M for EGF. HGF was additive with EGF in stimulating [3H] thymidine incorporation. In addition to rabbit proximal tubule cells, HGF increased proliferation in a cultured mouse proximal tubule cell line, MCT, and in rat glomerular epithelial cells. In contrast, HGF did not stimulate proliferation of either rat mesangial cells or a rat aortic smooth muscle cell line, A7r5. The HGF receptor is the product of the c-met proto-oncogene. C-met mRNA was detected in total kidney and in cultured proximal tubule cells but was not detected in cultured mesangial cells. In contrast, HGF mRNA was detected in mesangial cells but not in cultured proximal tubule cells. Preincubation of rabbit proximal tubule cells with the tyrosine kinase inhibitor, genistein (50 microM), prevented HGF-stimulation of [3H] thymidine incorporation. In LiCl pretreated rabbit proximal tubule cells loaded with [3H] myoinositol, HGF increased total inositol phosphate release, measured by anion exchange chromatography (control: 2181 +/- 414 vs HGF: 2609 +/- 478 cpm/10(6) cells; n = 6; p < 0.05). Although genistein did not affect baseline phosphoinositide hydrolysis, it inhibited the HGF stimulation. Thus, HGF is mitogenic for cultured proximal tubule cells as well as glomerular epithelial cells. Inhibition of proliferation and PI turnover by genistein suggests that HGF's actions are mediated in part by tyrosine kinase activity. In mammalian kidney, HGF released from mesangial cells may serve as a paracrine activator of the adjacent epithelial cells.

Animals↗
PubMed1993

Effect of carnitine supplement to the dam on plasma carnitine concentration in the sucking foal.

The changes in carnitine in plasma and milk during the first 3 months of lactation were studied in 14 broodmares and their foals. Six of the mares (Group S) were given a supplement of 10 g carnitine split between the morning and evening feeds, starting 2 weeks before birth. At birth the plasma carnitine concentration in Group S mares was about twice that in Group NS mares (no supplement). In both groups the concentration initially declined in the days after birth. Whilst this trend was reversed in Group S mares, the concentration in Group NS mares remained at a reduced level for the remainder of the study. Milk concentrations declined continuously over the monitoring period in both groups. There was no apparent relationship between milk and plasma concentrations. Despite this the milk concentration tended to be higher in Group S than in Group NS mares although differences were not significant. There was an immediate drop in the plasma concentration in foals after birth which was reversed in foals of Group S mares but not in those of Group NS mares. There were no apparent side effects of carnitine supplementation.

Animal Feed↗