Combination therapy for the treatment of hepatitis C in the veteran population: higher than expected rates of therapy discontinuation.

AIM: To compare the efficacy of high-dose induction with standard dose interferon therapy for the treatment of chronic hepatitis C virus at the Dallas Veterans Affairs Medical Center. METHODS: Patients were randomized to receive 5 million units daily interferon-alpha2b for 4-weeks followed by 44-weeks for genotype 1 or 20 weeks for non-genotype 1 of standard dose therapy (3 million units three times a week) or standard dose therapy for total treatment duration. Daily weight-based ribavirin was used for entire therapy interval. RESULTS: Forty-five patients were enrolled in the trial with genotype 1 comprising 75.6% of the sample. Cirrhosis or bridging-fibrosis was present in 69% of the patients. Of the 29 liver biopsies available for Knodell scoring, 41% and 51% had scores of 6-10 and 11-15, respectively. Rates of sustained virological response did not differ significantly between the two treatment groups. Therapy type and/or early intervention for depression did not affect the rate of therapy discontinuation, which was 26.6%. CONCLUSION: The rate of sustained virological response was similar between the two treatment groups and higher than anticipated among patients with cirrhosis or bridging-fibrosis. The rate of therapy discontinuation was also higher than anticipated but was not attributable to therapy type or untreated depression.

Neuropeptide Y fails to normalize food intake in zinc-deficient rats.

Zinc deprivation results in decreased and cyclic food intake in rats. We determined the response of zinc-deprived rats to neuropeptide Y (NPY). In a preliminary experiment, rats were fed a low (-Zn; <1 mg/kg) or adequate zinc diet (+Zn; 100 mg/kg) for 4 days. NPY (5 or 10 microg) was then administered via an intracerebroventricular (ICV) cannula and food intake measured for 4 h. NPY stimulated food intake in all rats, but the difference in food intake due to zinc deprivation persisted. In a subsequent experiment, rats were fed the low zinc and adequate zinc diets for 4, 5 or 6 days. Food intake was suppressed in rats fed the low zinc compared to the adequate zinc diet on all of these days. When NPY (10 microg) was administered at the onset of the light cycle, the food intake was approximately 2.5-fold greater regardless of dietary zinc status, but the amount of food consumed by rats fed low zinc was approximately one-half the quantity consumed by NPY-stimulated zinc-adequate rats. NPY administered at the onset of dark failed to stimulate food intake in either dietary group although the total intake difference due to zinc status persisted. ICV administration of 5 nmol of zinc prior to NPY injection failed to correct the food intake response of the zinc-deficient rats. We conclude that the basis of the reduced food intake of zinc-deficient rats does not relate to NPY quantity or release, or to impairment of its signal transduction. There appears to be another undefined factor that limits food intake in zinc deficiency.

Zinc Deprivation and the Nervous System.

This review is concerned with the role of zinc in the function of the nervous system with emphasis on the effects of dietary zinc deprivation. Zinc deficiency leads to several pathological signs, some of which occur within a few days in growing animals deprived of zinc. Depletion eventually leads progressively to more severe behavioral changes, to abnormal stance, and to peripheral neuropathy. The nervous system contains many essential zinc metalloproteins that are highly stable, i.e. have high zinc association constants. These proteins, which include enzymes, transcription factors and storage proteins, do not become limiting during incipient zinc deficiency. It is likely that other, yet unidentified, zinc dependent proteins become limiting within a few days after animals are deprived of dietary zinc and lead to behavioral changes such as decreased and cyclic feeding. One candidate for the first limiting zinc component of nerve tissue is the "chelatable" zinc pool. Another is the plasma membrane zinc pool; the latter pool is decreased in zinc deficient rat erythrocytes, leading to oxidation of protein thiol groups and malfunction of that membrane. While the defective biochemical mechanism(s) that leads to the signs of deficiency is not entirely clear, there is evidence that synaptosomes from zinc deficient guinea pigs do not take up calcium normally when stimulated. It is postulated that the most vulnerable zinc pool serves a protective role against oxidative damage to plasma membrane proteins including the proteins involved with calcium channels. If so, the first limiting role of zinc is protective rather than catalytic or that of a neurotransmitter.

Decreased plasma membrane thiol concentration is associated with increased osmotic fragility of erythrocytes in zinc-deficient rats.

Zinc deficiency leads to pathological signs that are related to impaired function of plasma membrane proteins. The purpose of this study was to assess the effect of dietary zinc status on the sulfhydryl (SH) content of erythrocyte plasma membranes and erythrocyte function. Three experiments were performed. In the first, immature male rats were fed for 21 d either a low-zinc (<1.0 mg/kg) diet free choice (-ZnAL), an adequate-zinc (100 mg/kg) diet free choice (+ZnAL), or the adequate-zinc diet limited to the intake of -ZnAL pair-mates (+ZnPF). Tail blood was sampled to measure osmotic fragility and SH concentration of erythrocyte membrane proteins. The zinc-deficient rats were then repleted for 2 d and erythrocytes assayed for fragility and SH content. In the second experiment blood was sampled at 3-d intervals to determine the time course of change in fragility and SH concentration. In the third experiment the SH concentration of erythrocyte band 3 protein and the binding of zinc to isolated plasma membranes were measured. SH concentration decreased from approximately 75 nmol/mg protein to 68 nmol/mg protein during 21 d of depletion and returned to control level within 2 d of repletion. There was an inverse relationship between osmotic fragility and SH concentration of erythrocyte membrane proteins. Maximal decrease in SH occurred within 6 d of consuming the low-zinc diet. The SH content of band 3 protein isolated from deficient rats was also significantly lower than that of pair-fed controls (45 vs. 51 nmol/mg protein). The zinc-binding affinity of plasma membrane proteins tended to be decreased by zinc deficiency. In summary, low-zinc status lowers the plasma membrane SH concentration, and the decreased reducing potential is inversely related to osmotic fragility, and presumably, with impaired volume recovery of erythrocytes.

Zinc deprivation of murine 3T3 cells by use of diethylenetrinitrilopentaacetate impairs DNA synthesis upon stimulation with insulin-like growth factor-1 (IGF-1).

Growth failure in zinc-deficient animals is associated with decreased DNA synthesis; zinc deprivation of 3T3 cells, by use of diethylenetrinitrilopentaacetate (DTPA), impairs thymidine incorporation when the cells are stimulated with fetal bovine serum (FBS). The purpose of this study was to determine the step of cell cycle progression that is affected by zinc deprivation. Swiss murine 3T3 cells were cultured for 3 d in complete media and then for 2 d in low serum media. Cells were then placed in serum-free media and stimulated in sequence with platelet-derived growth factor (PDGF; 3 h), epidermal growth factor (EGF; 0.5 h) and insulin-like growth factor-I (IGF-I; 16 h). The combination of growth factors stimulated thymidine incorporation to the same extent as 10% FBS, and DTPA or EDTA (0.6 mmol/L) inhibited thymidine incorporation. Inhibition was prevented by addition of zinc, but not calcium, iron or cadmium (0.4 mmol/L). When DTPA was present during all stages with no addition of zinc, or zinc added during the competency-priming (PDGF and EGF) step, the IGF-I step, or both steps, the zinc effect occurred at the IGF-I step. Zinc addition 4 h before the measurement of thymidine incorporation had no ameliorative effect, but the presence of zinc during the prior 12 h increased incorporation. Thus zinc exerts its major effect on DNA synthesis during the IGF-I stimulatory phase of the cell cycle. The total zinc concentration of 3T3 cells treated with DTPA for 16 h was not different from that of untreated cells; hence only a small compartment of the cell is affected by DTPA.

Chelation of extracellular zinc inhibits proliferation in 3T3 cells independent of insulin-like growth factor-I receptor expression.

Depletion of zinc inhibits growth in animals and proliferation of cultured cells. Additionally, zinc can serve as an antioxidant protecting many compounds, including proteins, from oxidation. Regulation of cell division also involves insulin-like growth factor type I (IGF-I) and its receptor, especially during late G1 phase, allowing progression of the cell to S phase with subsequent DNA synthesis. We examined the effects of zinc depletion from the culture media of Swiss 3T3 cells on the cell cycle and IGF-I receptor expression. Cells were exposed to reduced fetal bovine serum concentrations to induce growth arrest, then returned to normal fetal bovine serum concentrations with the divalent cation chelator diethylenetriamine pentaacetic acid. Reducing the fetal bovine serum concentration did not induce quiescence in the cells as previously suggested. Zinc depletion reduced the proliferative fraction (S and G2/M phases) of the cell cycle. The addition of glutathione to the zinc-depleted media partially returned the proliferative fraction to the control level. Fetal bovine serum deprivation reduced IGF-I receptor expression whereas the absence of zinc had little effect on receptor expression. We conclude that depletion of zinc from culture media inhibits 3T3 cell proliferation independent of insulin-like growth factor-I receptor expression, and part of this inhibition is due to the antioxidant capacity of this divalent cation.

Reduced food intake in zinc deficient rats is normalized by megestrol acetate but not by insulin-like growth factor-I.

Zinc deficiency in rats results in impaired growth accompanied by decreased and cyclic food intake. These signs are associated with decreased plasma insulin-like growth factor-I (IGF-I), a major mediator of growth. The purpose of this study was to determine the relationship between decreased plasma IGF-I and the impairment of appetite and growth in zinc deficiency. Immature male rats were fed free choice a low zinc (<1 mg/kg) diet (-Zn) or a zinc adequate (100 mg/kg) control diet (+Zn). Plasma IGF-I concentrations were normalized in zinc-deficient rats by the following two methods: osmotic pump infusion of IGF-I (2.4 mg/kg body weight daily) and oral administration (50 mg/kg body weight twice daily) of the synthetic progestin, megestrol acetate (MA). Infusion of IGF-I for 8 d sustained plasma IGF-I concentrations in zinc-deficient rats at control levels but had no effect on either food intake or growth rate. MA administration for 8 d maintained the plasma IGF-I of deficient rats and significantly increased food intake. The early aspects of cyclic food intake were eliminated, and, after a few days, food intake of deficient rats given MA was not different than that of controls. MA increased food intake and fat deposition regardless of zinc status, but it had no effect on the growth rate of deficient rats. MA significantly decreased body weight of controls, uncoupling energy intake and gain. The results suggest that reduced food intake precedes the decreased plasma IGF-I concentration and that IGF-I is not responsible for the decreased growth and food intake of zinc-deficient rats. The appetite and growth impairment of zinc-deficient rats may arise from disrupted function of IGF-I receptors in the brain and peripheral tissues, but not from low circulating levels of IGF-I.

Zinc deficiency decreases the concentration of N-methyl-D-aspartate receptors in guinea pig cortical synaptic membranes.

Zinc deficiency in guinea pigs decreases glutamate-stimulated calcium uptake in cortical synaptosomes. Glutamate not only stimulates calcium uptake but also potentiates the binding of the drug dizocilpine (MK-801) to an internal site of the N-methyl-D-aspartate receptor/calcium channel, a subtype of the glutamate receptor. The purpose of this study was to determine whether the effect of zinc deficiency on calcium uptake by glutamate-stimulated synaptosomes is related to N-methyl-D-aspartate receptor number or function, as measured by MK-801 binding. Immature guinea pigs consumed a low zinc (< 1 mg/kg) diet ad libitum or an adequate zinc (100 mg/kg) diet, either ad libitum or restricted to maintain weight similar to that of the low zinc animals. Binding of MK-801 to cortical membranes was measured first in the presence of saturating concentrations of glutamate or N-methyl-D-aspartate in combination with glycine. Zinc deficiency significantly reduced the concentration of MK-801 binding sites (20%) regardless of the potentiating agonist used, but had no effect on binding affinity. The binding of MK-801 in response to 1, 10 and 100 mumol/L glycine, in the presence of 100 nmol/L glutamate, was then measured and found to be significantly reduced (12%). The results suggest that zinc deficiency decreases the number of functional N-methyl-D-aspartate receptor/channels in cortical membranes, probably because of impaired channel opening.

Low zinc status in guinea pigs impairs calcium uptake by brain synaptosomes.

Zinc deficiency results in defective central nervous system function and in peripheral neuropathy. Calcium serves as second messenger in both pre- and postsynaptic membranes. Presynaptic uptake of calcium occurs via voltage-gated channels, whereas postsynaptic uptake occurs by way of a glutamate-activated channel, the N-methyl-D-aspartate (NMDA) receptor-channel. This study was designed to determine the effect of zinc status on calcium uptake by synaptic membranes prepared from guinea pigs deprived of zinc. Within each group of three guinea pigs, one animal was allowed to consume a low zinc (< 1 mg/kg) diet ad libitum (-ZN), one an adequate zinc (100 mg/kg) diet ad libitum (+AL), and one the adequate zinc diet restricted (+RF). When the -ZN guinea pig within a group developed clinical signs of deficiency, synaptosomes were prepared from brain cortices and calcium uptake measured by use of 45Ca. Both high potassium- and glutamate-stimulated calcium uptakes by synaptosomes from zinc-deficient guinea pigs were significantly lower than those of controls, with the glutamate-stimulated uptake 40% lower. In vitro addition of either magnesium or zinc resulted in lower uptake in synaptosomes from all dietary groups. Regardless of in vitro conditions, calcium uptake was impaired by zinc deficiency. The impaired function of calcium channels may explain the neurological disturbances observed in zinc-deficient animals.

Effect of zinc deficiency on enzyme activities in rat and pig erythrocyte membranes.

There is need for a reliable index of zinc status in humans. Considering the importance of zinc in membrane function, activities of erythrocyte membrane enzymes have been measured in animals of low and normal zinc status as possible indices. Immature rats and neonatal pigs were fed low and adequate zinc diets; the latter was fed both ad libitum and restricted so as to control for food intake effects. Low rates of gain and plasma zinc concentrations demonstrated that animals fed the low zinc diets were of low zinc status. Erythrocyte membranes were prepared and assayed for Na,K-ATPase, 5'-nucleotidase, and calcium-ATPase activities. Na,K-ATPase activity was not affected by zinc status, but 5'-nucleotidase was significantly lower in deficient animals of both species than in controls, whose food intake was restricted to maintain comparable weight (2.76 vs 3.94 nmol/hr/mg of protein in rats and 60.5 vs 119 in pigs). The basal calcium-ATPase activities were also decreased by low zinc status in both species. Addition of calmodulin in vitro stimulated activity two-fold to four-fold and resulted in the same maximal activities for all treatments. The results show that erythrocyte membrane 5'-nucleotidase activity is an index of zinc status in these species. It is suggested that the decreased membrane calcium-ATPase activity in zinc deficiency is caused by a defect in calmodulin metabolism.

Impaired hemostasis and platelet function in rats fed low zinc diets based on egg white protein.

Zinc-deficient rats exhibit impaired hemostasis, a pathological sign related to defective platelet function. The original observation was made in rats fed a low zinc diet based on soy protein. The present study was designed to test the effect of feeding a low zinc, egg white-based diet on bleeding time and platelet aggregation. The reversibility of the defect and the response of washed platelets from rats fed low zinc (less than 1 mg/kg) and control (100 mg/kg) diets were also assessed. Immature male rats were fed the respective diets for 11 d, the controls being both ad libitum- and pair-fed. To test reversibility, rats depleted for 11 d were then fed the control diet for an additional 7 d. Saline bleeding time was increased, and aggregation of platelet-rich plasma (PRP) from rats of low zinc status was impaired. The rate of the secondary phase of aggregation was significantly less than that of controls, but it was not different from that of controls after 7 d of zinc repletion. Aggregation of washed platelets was also impaired by low zinc status, showing that the defect is associated with the platelet, not the plasma. Egg white as a source of protein in the low zinc diet resulted in abnormal hemostasis and platelet aggregation, including a defective response of washed platelets.

Zinc status and peripheral nerve function in guinea pigs.

Guinea pigs fed a diet low in zinc develop clinical signs of apparent neurological origin. The signs include abnormal posture and locomotion as well as hypersensitivity to touch. In this study, electrophysiological and biochemical measurements were made on sciatic nerves from zinc-deficient and repleted animals as well as on controls fed either ad libitum or restricted to maintain weight comparable to those consuming the deficient diet. Both in vivo and in vitro measurements showed decreased motor nerve conduction velocity (NCV) in nerves of deficient animals. A longitudinal study showed excellent correlation of NCV and severity of clinical signs. Nerves from zinc-deficient guinea pigs had decreased Na,K-ATPase activity, but the number of sodium channels, as determined by saxitoxin binding, was not affected. It was concluded that the clinical signs of neuropathy in zinc deficiency are associated with impaired NCV and decreased Na,K-ATPase activity of peripheral nerves. The zinc-deficient guinea pig provides a useful model to study the biochemical defect in a peripheral neuropathy.

Zinc deficiency and peripheral neuropathy in chicks.

Zinc-deficient chicks develop an arthritic-like neuromuscular disorder. They walk with a stilted gait and tend to remain in a squat position, bearing little weight on the legs. The purpose of this study was to determine the basis of the syndrome by making electrophysiologic measurements of nerve function. Chicks were fed low zinc (6 mg/kg) and zinc-adequate (50 mg/kg) diets, the latter ad libitum and pair-fed. At the end of 3 weeks, sciatic nerve function was determined in vivo by use of an electrodiagnostic system. Motor nerve conduction velocity was significantly lower in chicks fed the low zinc than in those fed the zinc-adequate diet. Zinc repletion of the 2-week depleted chicks was achieved by feeding the adequate diet for 2 weeks. Repletion for this period cured clinical signs and restored nerve conduction velocity to normal, but reversal did not occur within 1 week. It was concluded that the abnormal posture and locomotion of zinc deficiency are associated with peripheral neuropathy.

Production and reversal of the neuromuscular pathology and related signs of zinc deficiency in guinea pigs.

The purposes of this study were to use vocalization and posture as indices of the neuromuscular pathology that develops in zinc-deficient guinea pigs and to determine the rate of depletion before and of repletion after intraperitoneal (IP) zinc therapy. When severe signs of deficiency developed, tissues were analyzed for their zinc content to assess sites of depletable and mobilizable zinc stores. Weanling guinea pigs were fed low (less than 1 mg/kg) and adequate (100 mg/kg) zinc diets based on isolated soybean protein or autoclaved egg white. The first signs (stage 1) of vocalization due to handling, abnormal posture and skin lesions developed after approximately 4 wk. Severe (stage 3) signs followed after 5-6 wk. A single IP dose of ZnSO4 (50 mumol/kg) caused remission of signs within 4-5 d following which all signs regressed to stage 3 within 7 d. Analysis of 15 tissues from severely deficient guinea pigs showed that only plasma and bone had significantly lower zinc concentrations than tissues from comparable age-related controls fed a zinc-adequate diet. It seems unlikely that major soft tissues, such as muscle, brain, liver and skin, serve as mobilizable stores of zinc for other critical metabolic functions. Bone zinc is slowly mobilized but at a rate insufficient to maintain health or even life.

Zinc deficiency increases the osmotic fragility of rat erythrocytes.

Zinc deficiency in rats causes increased osmotic fragility of their erythrocytes. This study was designed to determine the relationship of food intake and dietary sulfur amino acid level to the effect of low zinc status on fragility. Immature rats were fed for a 3-wk period a low zinc diet (less than 1 mg/kg) based on isolated soybean protein or a similar control diet (100 mg Zn/kg diet) supplied either ad libitum or by pair feeding. Fragility was measured by the degree of hemolysis in hypotonic saline solutions. In the first experiment, zinc deficiency resulted in higher fragility than in ad libitum controls; pair-fed controls were intermediate and not different from either. Experiment 2 included two levels of methionine, 0.4 and 0.9%, and two of zinc, 0 and 100 mg Zn/kg diet. At the 0.4%, but not at the 0.9% methionine level, hemolysis of red blood cells from the zinc-deficient rats was significantly greater than those from either pair-fed or ad libitum controls. Repletion for 1 or 2 d completely alleviated the increased fragility, but in vitro addition of zinc had no effect. Restricted intake of the zinc-adequate diet reversed the fragility within 1 d as readily as did ad libitum intake. Thus, the osmotic fragility induced by zinc deficiency was prevented by high sulfur amino acid intake and was readily reversed by dietary zinc. It is postulated that extracellular or membrane-bound zinc protects a component of the membrane that is essential to its function, and that reversal of the defect requires an in vivo metabolic process.

Zinc deficiency in rats reduces the vasodilation response to bradykinin and prostacyclin.

The objective of this study was to determine the effect of zinc deficiency on the blood pressure response of rats to intravenously administered doses of two chemically distinct vasodilators, bradykinin (BK) and prostacyclin (PGI2). Immature albino rats were fed a low zinc (less than 1 ppm) diet based on soybean protein, a similar control diet (100 ppm) ad libitum or the control diet, pair-fed. After 3 wk, the carotid arteries were catheterized for blood pressure measurement and graded doses of the vasodilators were injected as a bolus into a branch of the femoral vein. In one experiment graded doses (0-8 micrograms/kg body wt) of Bk and in another graded doses (0-400 ng/kg body wt) of PGI2 were injected. In a third experiment the PGI2 doses ranged from 0 to 1000 ng/kg and, in addition to the 3-wk depletion group, two groups were repleted by feeding the control diet for 1 or 4 d. The low zinc diet induced zinc deficiency as evidenced by low rates of gain and low plasma zinc levels and significantly (P less than 0.05) decreased the blood pressure response (Emax) to both BK and PGI2. Pair feeding had no significant effect on the response; the Emax values as determined by double-reciprocal plots of the data were unchanged by the quantity of control diet consumed. Short-term repletion did not restore the PGI2 response to control values. The data are interpreted as suggesting that zinc deficiency impairs the function of vasodilator receptors in the rat vasculature.

Effect of positional loading of three removable partial denture clasp assemblies on movement of abutment teeth.

Tests demonstrated that the degree of movement of the abutment tooth for a distal-extension base using three different clasp designs was a function of the location of loading. Thus, the null hypothesis, there is no significant difference in the degree of movement of the abutment tooth around the axis of rotation as a function of the position of loading, has been demonstrated to be invalid based on the data obtained in this investigation.

Movement of three removable partial denture clasp assemblies under occlusal loading.

This investigation indicates that the design of the clasp assembly affects the magnitude of movement of the abutment tooth adjacent to an extension base but generally does not affect the direction of movement. Some clasp assemblies (distal rest) moved more than other clasp assemblies (wrought wire or I-bar), and hence, the abutment tooth moved more. The design of the clasp assembly did not seem to affect the magnitude of movement in the apical direction.