Search PubMed⌕ Search

Biomedical subjects

J Gamble

Publications and source records attributed to J Gamble.

At least 19 recordsLinked to original sources
PubMed2002

The effect of chronic insulin delivery via the intraperitoneal versus the subcutaneous route on hepatic triglyceride secretion rate in streptozotocin diabetic rats.

Chronic intraperitoneal or subcutaneous insulin administration increases triglyceride secretion rate (TGSR) in normal rats. We wished to determine the effect of this treatment on TGSR and the hepatic lipogenic enzymes acetyl-CoA carboxylase (ACC) and fatty acid synthase (FAS) in diabetic rats. Streptozotocin-diabetic rats, untreated (D), diabetic rats treated with insulin (3 U/day for 21 days) intraperitoneally (IP) or subcutaneously (SC) and non-diabetic rats (N) were studied. TGSR was determined using Triton WR-1339. Fasting glucose and triglyceride levels, high in D, were normalized by insulin treatment regardless of route. Peripheral insulin levels were lowest in D and highest in SC, portal insulin levels were lowest in D and highest in IP. Non-esterified fatty acid levels were not elevated in D, presumably due to adipose tissue depletion. TGSR was reduced in D (P<0.05) and was normalized following insulin administration, regardless of route. ACC activity was normal, but FAS was decreased in D (P<0.05). ACC and FAS were normal in both IP and SC. Thus, in streptozotocin-diabetic rats, chronic intraperitoneal or subcutaneous insulin treatment increases TGSR and FAS activity from their low levels in insulin-deficient rats to levels equal to but not higher than those in normal rats.

Acetyl-CoA Carboxylase↗
PubMed2001

Noninvasive measurement of microvascular leakage in patients with dengue hemorrhagic fever.

Dengue shock syndrome (DSS) is a potentially lethal complication of dengue virus infection associated with hypotension and leakage of plasma water into the extravascular space. To determine whether the underlying pathophysiology of DSS is distinct from that in milder forms of the disease, we assessed microvascular permeability, by use of strain gauge plethysmography, in Vietnamese children with DSS (n=19), or dengue hemorrhagic fever (DHF) without shock (n=16), and in healthy control children (n=15). At admission and after fluid resuscitation, the mean coefficient of microvascular permeability (K(f)) for the patients with dengue was approximately 50% higher than that for the control patients (P=.02). There was no significant difference in K(f) between the 2 groups of patients with dengue; this suggests the same underlying pathophysiology. We hypothesize that in patients with DSS, the fluctuations in K(f) are larger than those in patients with DHF, which leads to short-lived peaks of markedly increased microvascular permeability and consequent hemodynamic shock.

Adolescent↗
PubMed2001

A longitudinal study of resting peripheral blood flow in normal pregnancy and pregnancies complicated by chronic hypertension and pre-eclampsia.

OBJECTIVES: To investigate the hypothesis that reduced resting tissue blood flow precedes the clinical onset of pre-eclampsia in women at risk of the disease. METHODS: We used venous occlusion plethysmography to compare resting calf muscle blood flow in 18 normal pregnant controls, 18 pregnant women with chronic hypertension, and 23 pregnant women at increased risk of developing pre-eclampsia. Calf blood flow was measured at 16, 20, 24, 28, 32 and 36 weeks of gestation. RESULTS: Blood flow increased with gestation in normal pregnancy (P = 0.004) and chronic hypertension (P = 0.006), but not in the 'at risk' women who did not develop pre-eclampsia (P = 0.36). In contrast, blood flow decreased significantly in eight out of the 23 women 'at risk', who developed pre-eclampsia (P < 0.00001, ANOVA). The decrease in flow preceded the clinical diagnosis of the pre-eclampsia by several weeks. Moreover, a significant inverse correlation was observed between resting blood flow and plasma uric acid concentrations (r = -0.86, P = 0.03) in the women that developed pre-eclampsia. CONCLUSIONS: We have shown that reduced resting blood flow precedes the clinical onset of pre-eclampsia independently of hypertension per se. These findings support the notion that impaired tissue blood flow may be involved at an early stage in the pathophysiology of the disease.

Chronic Disease↗
PubMed2001

Lack of effect of serum amyloid A (SAA) on the ability of high-density lipoproteins to inhibit endothelial cell adhesion molecule expression.

Studies have been conducted to determine whether the ability of high density lipoproteins (HDL) to inhibit the cytokine-induced expression of vascular cell adhesion molecule-1 (VCAM-1) in endothelial cells is altered by the presence in HDL of the acute phase reactant, serum amyloid-A (SAA). Preparations of HDL(3) were isolated on two separate occasions from the plasma of each of 19 patients: the first was collected before and the second 3 days after undergoing coronary artery bypass graft surgery. Whereas the preoperative HDL(3) sample contained no SAA, in the postoperative sample SAA accounted for an average of 42% of the HDL(3) protein. The preoperative HDL(3) and postoperative, SAA-enriched HDL(3) were identical in terms of their ability to inhibit the tumour necrosis factor-alpha (TNF-alpha)-induced expression of VCAM-1 in human umbilical vein endothelial cells (HUVECs). To assess the effect of having an even greater SAA enrichment of HDL(3), samples of HDL(3) were incubated with purified SAA, which displaced almost all of the apoAI and about 40% of the apoAII from the HDL(3). This in vitro SAA-enriched HDL(3) inhibited the TNF-alpha-induced expression of VCAM-1 in HUVECs in a concentration dependent manner, which was identical to that of the unmodified HDL(3). The presence of SAA did not alter the cell-surface binding of HDL(3) to endothelial cells. It has been concluded that the presence of SAA in HDL has no effect on the ability of these lipoproteins either to inhibit the expression of VCAM-1 in endothelial cells or to bind to proteins on the endothelial cell surface.

Apolipoprotein A-I↗
PubMed2001

Modifications of microvascular filtration capacity in human limbs by training and electrical stimulation.

This study investigated whether an increase in microvascular surface area as a result of endurance training, which increases human skeletal muscle capillarity, would translate to greater capacity for fluid filtration compared with strength training, which does not affect capillary supply. Values for filtration capacity, Kf, derived from the slope of calf volume change, Jv, measured by venous occlusion plethysmography, against cuff pressure during a protocol of small cumulative pressure steps, were significantly higher in endurance athletes (5.78 +/- 0.88 mL min(-1) 100 mL(-1) mmHg(-1) x 10(-3), P < 0.05) than controls (3.38 +/- 0.32 mL min(-1) 100 mL(-1) mmHg(-1) x 10(-3) whereas strength-trained athletes had values similar to control (4.08 +/- 0.56 mL min(-1) 100 mL(-1) mmHg(-1) x 10(-3), ns), suggesting that surface area is important. However, when sedentary subjects underwent either a 4-week unilateral dynamic plantarflexion training programme (70% peak power, 20 min day(-1), 5 days week(-1) or a calf muscle electrical stimulation programme (8 Hz, 3 x 20 min day(-1), 5 days week(-1), neither of which caused limb blood flow to alter after training nor would be expected to increase capillarity, only the stimulation group showed a significant increase in Kf (6.68 +/- 0.62 mL min(-1) 100 mL(-1) mmHg(-1) x 10(-3) post-training vs. 3.38 +/- 0.38 pre-training, P < 0.05). This may be because stimulation enhances perfusion preferentially to glycolytic fibres, or maintains high levels of vascular endothelial growth factor (VEGF) or changes lymph clearance.

Adaptation, Physiological↗
PubMed2001

Changes in microvascular fluid filtration capacity during 120 days of 6 degrees head-down tilt.

We used venous congestion strain gauge plethysmography (VCP) to measure the changes in fluid filtration capacity (K(f)), isovolumetric venous pressure (Pv(i)), and blood flow in six volunteers before, on the 118th day (D118) of head-down tilt (HDT), and 2 days after remobilization (Post). We hypothesized that 120 days of HDT cause significant micro- and macrovascular changes. We observed a significant increase in K(f) from 3.6 +/- 0.4 x 10(-3) to 5.7 +/- 0.9 x 10(-3) ml. min(-1). 100 ml(-1). mmHg(-1) (+51.4%; P < 0.003), which returned to pretilt values (4.0 + 0.4 x 10(-3) ml. min(-1). 100 ml(-1). mmHg(-1)) after remobilization. Similarly, Pv(i) increased from 13.4 +/- 2.1 mmHg to 28.9 +/- 2.8 mmHg (+105.8%; P < 0.001) at D118 and was not significantly different at Post (12.4 +/- 2.6 mmHg). Blood flow decreased significantly from 2.3 +/- 0.3 to 1.3 +/- 0.2 ml. min(-1). 100 ml tissue(-1) at D118 and was found elevated to 3.4 +/- 0.7 ml. min(-1). 100 ml tissue(-1) at Post. We believe that the increased K(f) is caused by a higher microvascular water permeability. Because this may result in edema formation, it could contribute to the alterations in fluid homeostasis after exposure to microgravity.

Adult↗
PubMed2000

Development of T-leukaemias in CD45 tyrosine phosphatase-deficient mutant lck mice.

The CD45 tyrosine phosphatase lowers T-cell antigen receptor signalling thresholds by its positive actions on p56(lck) tyrosine kinase function. We now show that mice expressing active lck(F505) at non-oncogenic levels develop aggressive thymic lymphomas on a CD45(-/-) background. CD45 suppresses the tumorigenic potential of the kinase by dephosphorylation of the Tyr394 autophosphorylation site. In CD45(-/-) thymocytes the kinase is switched to a hyperactive oncogenic state, resulting in increased resistance to apoptosis. Transformation occurs in early CD4(-)CD8(-) thymocytes during the process of TCR-beta chain rearrangement by a recombinase-independent mechanism. Our findings represent the first example in which a tyrosine phosphatase in situ prevents the oncogenic actions of a SRC: family tyrosine kinase.

Animals↗
PubMed2000

Altered peripheral vascular response of women with and without pelvic pain due to congestion.

OBJECTIVE: To test the hypothesis that women with pelvic venous congestion have a reduction of reactivity of their peripheral circulation. DESIGN: Comparison was made between 20 women with chronic pelvic pain due to congestion and a control group of 15 pain-free women matched for age, parity and body weight. A comparison of these results was made with those from six postmenopausal women taking hormone replacement therapy. METHODS: Study and control groups were investigated during the mid-follicular phase of the menstrual cycle (days 5-9) and the mid-luteal phase (days 19-23). The study group was also investigated during the fifth month of treatment with suppression of ovarian activity with leuprorelin or medroxyprogesterone acetate or six months after hysterectomy and bilateral salpingo-oophorectomy. Head-up tilt sufficient to increase intra-vascular pressure in the toe by a standard 40 mmHg was used as a means of raising venous pressure in the lower limb. Skin capillary red blood cell velocity (flux) was measured using a laser Doppler flow probe placed over the pulp of the big toe. Heart rate and blood pressure were also recorded. The change in skin blood flow following head-up tilt was expressed as a percentage of baseline flow in the supine position. MAIN OUTCOME MEASURES: Percentage change in skin red blood cell flux, heart rate and blood pressure in response to 40 degrees head-up tilt. RESULTS: In the control group the median response to head-up tilt in the follicular phase was one of a reduction in flux, whereas in the luteal phase it was more variable ranging from an increase to a decrease in flux. The responses in the pelvic congestion group in both the follicular and luteal phases were similar to those of the control group in the luteal phase. A small but significant increase in heart rate in response to tilt in the pelvic pain group, compared with the control group, was interpreted as being due to a fall in venous return. Treatment of the pelvic congestion group by medical suppression of ovarian activity or total hysterectomy with bilateral salpingo-oophorectomy resulted in a significant change in response to head-up tilt from the variable type of luteal response to one of a more constant reduction in flux, similar to that of the control group in the follicular phase. A reduction in flux was also found consistently in postmenopausal women. CONCLUSION: The study confirms the hypothesis that women with pelvic pain due to congestion show a change in peripheral vascular reactivity which returns to normal after suppression of ovarian activity. It seems likely that some alteration of normal ovarian function is responsible for the observed changes in peripheral blood flow in response to a rise in venous pressure in women with pelvic congestion.

Blood Flow Velocity↗
PubMed2000

Description and validation of a novel liquid metal-free device for venous congestion plethysmography.

We present a newly developed electromechanical sensor with automated calibration for strain-gauge plethysmography (filtrass) and compare it to a conventional mercury-in-Silastic strain-gauge plethysmograph (MSG). Fluid filtration capacity (K(f)) and isovolumetric venous pressure (Piv) of the limb were assessed noninvasively with both devices in 29 healthy volunteers. We found significantly higher K(f) and Piv values with MSG [4.6 +/- 2.0 x 10(-3) ml. min(-1). mmHg(-1). 100 ml tissue(-1) (K(f) units; K(f)U) and 21.2 +/- 8.1 mmHg for Pvi], than with filtrass, giving values of 3.1 +/- 0.8 K(f)U and 15.1 +/- 7.1 mmHg. Because K(f) and Piv are profoundly influenced by the calibration, we investigated the quality of the calibration signal and its impact on the obtained values. We could show that the reproducibility of repeated calibrations was higher with filtrass (58% lower mean +/- SD). The data were grouped according to the quality of calibration, and we found no significant difference in K(f) and Piv between filtrass (3.0 +/- 0.7 K(f)U and 15.9 +/- 6.9 mmHg, respectively) and MSG with good calibration signal (3.3 +/- 0. 8 K(f)U and 18.6 +/- 7.1 mmHg, respectively; no significant difference). However, we obtained significantly higher MSG values (5. 6 +/- 2.0 K(f)U and 23.1 +/- 8.4 mmHg, respectively; P < 0.001) in the group with a bad calibration signal. We suggest that the filtrass sensor, which performs an automatic, standardized calibration procedure and shows a linear signal response to stretch, gives highly reproducible and reliable results and thus is more suitable for routine application.

Adult↗
PubMed2000

Age-related changes in microvascular permeability: a significant factor in the susceptibility of children to shock?

During studies of the pathogenesis of dengue shock syndrome, a condition largely confined to childhood and characterized by a systemic increase in vascular permeability, we observed that healthy controls, age-matched to children with dengue shock syndrome, gave high values of filtration capacity (K(f)), a factor describing vascular permeability. We hypothesized that K(f) might be age dependent. Calf K(f) was studied in 89 healthy Vietnamese subjects aged 5 to 77 years. The K(f) was highest in the youngest children [7. 53 (1.96-15.46) K(f)U; median (range); where the units of K(f), K(f)U=ml.min(-1).100 ml(-1).mmHg(-1)]. Values were 3- to 4-fold lower towards the end of the second decade [4.69 (1.91-7.06) K(f)U]. Young mammals are known to have a larger microvascular surface area per unit volume of skeletal muscle than adults. During development the proportion of developing vessels is greater. Moreover, the novel microvessels are known to be more permeable to water and plasma proteins than when mature. These factors may explain why children more readily develop hypovolaemic shock than adults in dengue haemorrhagic fever and other conditions characterized by increased microvascular permeability.

Adolescent↗
PubMed2000

Vitamin C modifies the cardiovascular and microvascular responses to cigarette smoke inhalation in man.

Both neutrophil margination and increases in the non-invasively assessed parameter, isovolumetric venous congestion cuff pressure (Pv(i)), are symptomatic of some inflammatory diseases. Neutrophil margination occurs primarily, though not exclusively, at the post-capillary endothelial surface. The local haemodynamic changes resulting from margination may be responsible for the observed increases in Pv(i). Smoke inhalation has been shown in animal studies to cause an increase in post-capillary neutrophil margination by mechanisms that can be blocked by oral vitamin C administration. We looked for indices of a relationship between margination and Pv(i) in man, using cigarette smoke inhalation as a pathophysiological challenge. We also examined the effect of prophylactic vitamin C on the response. Smoke inhalation was associated with highly significant increases in both Pv(i) and heart rate. After vitamin C pre-treatment, no increase in Pv(i) was observed in response to the smoke inhalation; however, whilst heart rate still increased significantly, the duration of this response was attenuated. The results suggest that vitamin C affords protection against some of the cardiovascular and microvascular changes associated with cigarette smoke inhalation in man. They also support the notion that non-invasive assessment of changes in Pv(i) may provide a measurable index of systemic changes in inflammatory conditions.

Adult↗
PubMed2000

Resting peripheral blood flow in normal pregnancy and in pre-eclampsia.

Multiple organ dysfunction followed by end organ failure occurs in pre-eclampsia. While one would intuitively reason that one of the factors contributing to the end organ failure is poor nutritional blood flow, this has yet to be demonstrated. The aim of the present study was to determine whether changes in resting nutritional blood flow occur in pre-eclampsia. We used strain-gauge plethysmography to study calf blood flow in 19 women with pre-eclampsia, 13 normal pregnant women and 17 non-pregnant controls. We reasoned that, since the calf comprises mostly skeletal muscle, without anastomotic channels, blood flowing through this region would primarily reflect nutritive flow. Calf blood flow was significantly reduced in women with pre-eclampsia (1.95+/-0.9 ml.min(-1).100 ml(-1)) compared with normal pregnant (3.9+/-1.4 ml.min(-1).100 ml(-1)) and non-pregnant (3.8+/-1.0 ml.min(-1).100 ml(-1)) women (P=0.0004 and P=0.0005 respectively; ANOVA). Blood flow in pre-eclampsia was also correlated significantly with platelet count as an index of disease severity. In addition, there was a significant negative correlation between blood flow and systolic blood pressure (r=-0.69, P=0.004) in the women with pre-eclampsia. These findings support the hypothesis that nutritional blood flow is reduced in pre-eclampsia. We suggest that measurement of resting calf blood flow could give a non-invasive index of deterioration of nutritive blood flow to vital organs in pre-eclampsia.

Adult↗
PubMed1999

Microvascular fluid filtration capacity (Kf) assessed with cumulative small venous pressure steps and with various degrees of tilt.

UNLABELLED: Orthostatic dysregulation is a frequent phenomenon in pilots experiencing extreme G forces and after prolonged exposures to microgravity in cosmonauts. We used non-invasive venous congestion plethysmography (VCP) to study microcirculatory changes and the ability to activate peripheral and centrally mediated protective mechanisms during various degrees of tilt, which we used as an orthostatic challenge. METHOD: The study, which was approved by the local ethical committee of the Institute of Biomedical Problems, was performed on six healthy 20 - 26 years old male volunteers. We applied 6 - 8 cumulative small venous congestion pressure steps (8 mmHg) to the thigh and determined the fluid filtration capacity (Kf), the linear relationship between cuff pressure (Pcuff) and measured fluid filtration (Jv). We then measured the fluid filtration (Jv) response to varying cumulative degrees of tilt, starting at 0 degrees followed by head down -8 degrees -15 degrees, -30 degrees -15 degrees, -8 degrees, 0 degrees, and then head up 15 degrees, 30 degrees, 70 degrees, 30 degrees, 15 degrees and 0 degrees. Each tilt stage was sustained for 15 minutes. The change in hydrostatic load, at the level of the strain gauge, was determined by measuring the difference in vertical height between the right atrium and mid calf at the level of the gauge. Limb arterial blood flow was measured and lung function tests were performed before and after the small cumulative pressure step protocol, as well as at the end of each tilt step. RESULTS: No significant changes in blood pressure (BP) and heart rate (HR) occurred during the cumulative pressure step protocol. However, an increase in HR was observed during the tilt, but only reached significance at 30 degrees and 70 degrees. The mean Kf value measured with small cumulative pressure steps was 3.25 +/- 0.5 (10(-3) ml.100 ml tissue(-1) mmHg(-1) = KfU), which was significantly (p < 0.005) higher than the value obtained using tilt to increase the hydrostatic load (0.98 +/- 0.2 KfU). One subject had an unchanged Kf value and experienced vaso - vagal syncope following the imposition of 70 degrees tilt, with a heart rate < 35 / min and a systolic blood pressure of 60 mmHg. The measurement of blood flow in the limb showed no significant change during the tilt procedure. The lung function measurements revealed, that only expiratory reserve volume (ERV) did significantly increase during 35 degrees and 70 degrees head up tilt. All other parameters were unchanged. We suggest that the differences in these Kf values can be explained by the activation of both central and peripheral mediated changes in pre- capillary resistance in response to the tilt. We suspect, that the vaso - vagal syncope, experienced by one subject, reflected his inability to activate these protective reflex mechanisms, a situation that could be exacerbated by sub-clinical venous insufficiency. We believe that these results show that VCP is a useful tool for the determination of intolerance to orthostatic stress.

Adult↗
PubMed1999

[Preoperative changes in fluid filtration capacity in patients undergoing vascular surgery].

UNLABELLED: Patients undergoing major vascular surgery frequently require a substantial intraoperative fluid replacement to assure hemodynamic stability, which is in excess of the expected fluid requirements due to starving, blood and insensible losses. This leads to a positive fluid balance which can not be readily explained. METHOD: We have used venous congestion plethysmography (VCP) a non-invasive method for measurement of microvascular parameters in limbs to investigate the changes in microvascular permeability (FFK) and the balance of Starling forces of patients undergoing surgery for unilateral femoral artery reconstruction (FEM) under epidural anaesthesia or abdominal aortic aneurysm repair (AAA) under general anaesthesia. The control group consisted of patients scheduled for inguinal hernia repair or hand surgery under general anaesthesia. All patients were measured 24 hours pre-operatively, immediately after the induction of anaesthesia or completion of epidural anaesthesia and on the 1st, 5th and 10th postoperative day. The perioperative patient management followed a standard protocol and all patients with vascular disease were invasively monitored using indwelling arterial lines and central venous catheters. Continuous infusion of Ringers lactate and 6% Dextran 60 was sustained during the induction period. Each patient gave informed consent. RESULTS: Preoperatively we found no significant difference in the mean FFK-values of controls (4.1 +/- 0.4, ml.min-1 100 ml tissue-1 mmHg-1 x 10(-3) = FFKU), the AAA (3.6 +/- 0.3 FFKU) and FEM (4.2 +/- 0.3 FFKU). After induction of anaesthesia the mean FFK value in the controls fell to 3.5 +/- 0.5 FFKU (p = 0.07), whereas in the AAA patients we observed a significant increase to 4.7 +/- 0.2 FFKU (p < 0.005) and after epidural anaesthesia in FEM to 5.5 +/- 0.4 FFKU (p < 0.001) respectively. Those post anaesthetic FFK values where significantly higher in FEM and AAA than in the controls (p < 0.02). In AAA we found a significant positive correlation between the increase in FFK and the intraoperative fluid balance (r2 = 0.69, p < 0.01). No such correlation was found in controls and FEM. The postoperative values of FFK where unchanged in the control group, whereas a further increase was seen in both patient groups with vascular disease, with a maximum in AAA on the 1st postoperative day (to 5.4 +/- 0.4 FFKU mean both legs) and the 5th postoperative day in FEM (to 7.3 +/- 1.7 non-ischemic leg, 7.1 +/- 1.2 ischemic leg FFKU). In both groups normal FFK values where found on the 10th day after the operation. CONCLUSION: The data presented suggests an increase in extravascular fluid loss in patients undergoing vascular surgery, which becomes evident after the induction of general anaesthesia or completion of epidural anaesthesia. The positive correlation with the intraoperative fluid requirements may partially explain the often reported large intraoperative fluid requirements of patients undergoing AAA repair. The fact that the maximum change in fluid filtration capacity is found postoperatively may be explained by the additional effect of an ischemia/reperfusion injury in response to both the clamping an declamping of the artery and the increase in arterial blood flow to the limb due to the successful reconstruction of the blood vessel.

Adult↗
PubMed1999

B1 integrin activation inhibits in vitro tube formation: effects on cell migration, vacuole coalescence and lumen formation.

Human endothelial cells (EC), when plated onto gels of extracellular matrix proteins such as Matrigel or collagen form capillary tubes in a process thought to mimic angiogenesis. We have shown previously that the extent of tube formation and the phenotype of the lumen are regulated by integrins (Gamble et al 1993) and lumen formation occurs through a process of vacuolization, coalescence and ultimate directional fusion of these vacuoles with the plasma membrane (Meyer 1997 et al). We now show here that activation of beta1 integrins on endothelial cells inhibits tube formation. On collagen gels, endothelial cells treated with 31 activating antibody 8A2 failed to migrate into the gel and tube formation was inhibited. Although several integrins mediate EC attachment to collagen alpha2beta1 is the chief determinant of EC behaviour since a blocking antibody to (alpha2beta1 reversed the effect of 8A2. On Matrigel tube formation was also inhibited by 8A2 treatment although cell alignment and sprout formation was still evident. Electron microscopy revealed the organisation of normal numbers of cells into solid sprouts and the formation of small intracellular vacuoles suggesting that initial stages of tube formation including cell migration were unaffected. However, beta1 integrin activation inhibited the coalescence of these small vacuoles into larger vacuoles, the recruitment of more cells into the sprout and the subsequent formation of mature lumen. The inhibition of capillary tube formation by beta1 activation was time dependent and long lasting. The critical time for activation of the beta1 integrin was the initial 1-2h after plating in order to inhibit tube formation although once activated, the beta1 mediated inhibition on Matrigel was still evident 4 days later. Our results suggest that beta1 integrins are critical in capillary tube formation in at least two phases. beta1 integrins are essential for migration of EC through collagen gels. Independently, beta1 integrins, although not involved in initial vacuole formation, are involved in the process of vacuole coalescence and subsequent lumen formation since beta1 integrin activation inhibits these processes.

Antibodies, Monoclonal↗
PubMed1998

Functional domains of the alpha1 catalytic subunit of the AMP-activated protein kinase.

The AMP-activated protein kinase is a heterotrimeric enzyme, important in cellular adaptation to the stress of nutrient starvation, hypoxia, increased ATP utilization, or heat shock. This mammalian enzyme is composed of a catalytic alpha subunit and noncatalytic beta and gamma subunits and is a member of a larger protein kinase family that includes the SNF1 kinase of Saccharomyces cerevisiae. In the present study, we have identified by truncation and site-directed mutagenesis several functional domains of the alpha1 catalytic subunit, which modulate its activity, subunit association, and protein turnover. C-terminal truncation of the 548-amino acid (aa) wild-type alpha1 protein to aa 312 or 392 abolishes the binding of the beta/gamma subunits and dramatically increases protein expression. The full-length wild-type alpha1 subunit is only minimally active in the absence of co-expressed beta/gamma, and alpha1(1-392) likewise has little activity. Further truncation to aa 312, however, is associated with a large increase in enzyme specific activity, thus revealing an autoinhibitory sequence between aa 313 and 392. alpha-1(1-312) still requires the phosphorylation of the activation loop Thr-172 for enzyme activity, yet is now independent of the allosteric activator, AMP. The increased levels of protein expression on transient transfection of either truncated alpha subunit cDNA are because of a decrease in enzyme turnover by pulse-chase analysis. Taken together, these data indicate that the alpha1 subunit of AMP-activated protein kinase contains several features that determine enzyme activity and stability. A constitutively active form of the kinase that does not require participation by the noncatalytic subunits provides a unique reagent for exploring the functions of AMP-activated protein kinase.

AMP-Activated Protein Kinases↗