Primate performance decrements following acute soman exposure: failure of chemical countermeasures

Three experiments are reported: 1) a feasibility study on using laboratory primates repeatedly in behavioral toxicity studies of organophosphate (OP) agents or of chemical countermeasures against OPs; 2) a study of the efficacy of pyridostigmine pretreatment and 2-PAM therapy; and 3) a study to determine the effects of these treatments on soman-induced cholinesterase (ChE) inhibition and its recovery. In rhesus monkeys, three repeated acute low-dose (2.1 to 2.8 micrograms/kg) soman exposures, separated by intervals > 5 weeks, did not change baseline compensatory tracking performance or the soman ED50. Atropine therapy (97 micrograms/kg) alone had no effect on soman ED50. Addition of pyridostigmine pretreatment (150 micrograms/kg) and 2-PAM therapy (17 mg/kg) to atropine therapy increased the soman ED50 for a performance decrement from 2.27 micrograms/kg to 2.58 micrograms/kg, an insignificant protective effect. At the soman ED50 for behavioral decrements, pyridostigmine pretreatment increased the inhibition of serum ChE observed immediately after soman exposure, but reduced the extent of permanent inhibition. The 2-PAM therapy reduced serum ChE inhibition from about 80% to less than 70%. These effects on the time course of ChE inhibition following soman exposure appear to combine additively. These chemical countermeasures do not prevent soman-induced performance decrements, even though they are effective in protecting lives after much higher doses. The soman doses used produce only small, transient performance decrements; animals so exposed can, thus, be used repeatedly in such studies.     

Early postinfection antiviral treatment reduces viral load and prevents CD4+ cell decline in HIV type 2-infected macaques

Reports of significant reductions in plasma viral load by anti-HIV drugs have raised the possibility that antiviral therapy, if initiated sufficiently early, may result in sustained control of infection and prolonged clinical benefits. We evaluated the effects of intervention coincident with infection using an antiviral nucleoside, d4T, in Macaca nemestrina infected with a highly pathogenic isolate of HIV-2 (HIV-2[287]). Infection with this virus reproducibly results in high viremia and rapid CD4+ cell depletion, allowing a sensitive measurement of the treatment effect on viral load and clinical outcome. Compared to the control group, d4T-treated macaques showed significantly lower (2-3 log10) plasma- and cell-associated viral load. No CD4+ cell decline was observed in the treatment group while on therapy with d4T whereas CD4+ cells of control macaques declined from a preinfection mean of 32% of PBMCs to below 10%. Notably, when d4T treatment was withdrawn after 16 weeks, five of the six macaques continued to control viral load and have maintained normal CD4+ cell level for more than a year. These results demonstrate that early antiviral intervention, even of a limited duration, may constitute an important strategy against lentiviral-induced disease.     

Cyclical etidronate in the treatment of postmenopausal osteoporosis: efficacy and safety after seven years of treatment

PURPOSE: To determine the efficacy and safety of cyclical etidronate for up to 7 years in the treatment of postmenopausal osteoporosis and to examine the effects of discontinuing treatment after 2 or 5 years of therapy. PATIENTS AND METHODS: Patients were randomized at entry into the original study in 1986 to blinded treatment for 2 years with either a calcium (placebo) or an intermittent cyclical etidronate regimen, which most patients continued for a third year. Following this phase of the study, patients were enrolled into an open-label, follow-up study (years 4 and 5), during which all patients received cyclical etidronate treatment. In the present double-blind study (years 6 and 7), patients were rerandomized to receive intermittent cyclical therapy with either etidronate or placebo; all patients received calcium. The treatment regimen consisted of 400 mg/day etidronate or placebo for 14 days, followed by 76 days of elemental calcium (500 mg/day); this cycle was repeated approximately 4 times in each year. Of the 193 patients who continued in years 6 and 7 of the study, 93 were randomized to receive cyclical etidronate and 100 were randomized to receive calcium only. For purposes of efficacy analyses, patients were categorized by their total years of cumulative etidronate treatment (7, 5, 4, or 2 years). There were 51, 46, 42, and 54 patients in the 7-, 5-, 4-, and 2-year groups, respectively. Annual assessments included lumbar spine bone mineral density (BMD), as measured by densitometry, and vertebral radiographs. RESULTS: The groups receiving cyclical etidronate during this 2-year study period (7- and 4-year groups) had statistically significant mean percent increases in spinal BMD of 1.8% and 2.2%, respectively (P < 0.05) at the week 104 observation time. The 5- and 2-year groups, which did not receive etidronate during this period, had mean values of 1.4% and 0.2%, respectively (not significant) at week 104. In the 7-, 5-, 4-, and 2-year groups, the increases in spinal BMD at the end of 7 years were 7.6%, 8.6%, 8.1%, and 3.9%, respectively; these values were statistically significant for all groups compared with original baseline (year 0) (P < 0.05). BMD of the femur and wrist was maintained throughout the 7-year period. The incidence and rate of vertebral fractures were lowest in patients with the longest exposure to etidronate. Etidronate was well tolerated during the study, with low incidences of gastrointestinal side effects and nonvertebral fractures. CONCLUSIONS: Long-term cyclical etidronate is a safe, effective, and well-tolerated treatment for postmenopausal osteoporosis. Bone mass is maintained for at least 2 years after treatment with etidronate is stopped; however, further gains in spinal bone mass are seen in patients who continue therapy.     

The imidazoline I1 receptor agonist, moxonidine, inhibits insulin secretion from isolated rat islets of Langerhans

In order to study the pharmacology of the putative imidazoline receptor involved in stimulation of insulin secretion, the potent and selective imidazoline I1 receptor agonist, moxonidine, was employed. Surprisingly, this agent caused a rapid and complete inhibition of glucose-induced insulin secretion in isolated rat islets of Langerhans. This response was reversible upon removal of the compound but was only partially attenuated under conditions of complete alpha 2 blockade, suggesting that it did not derive entirely from the weak alpha 2-adrenoceptor agonist activity of moxonidine. Furthermore, the response could not be attributed to activation of imidazoline I1 receptors since it was not reproduced by a second potent imidazoline I1 receptor agonist, cimetidine, and could not be alleviated by the imidazoline I1 receptor antagonist efaroxan. The results confirm that the imidazoline receptor involved in control of insulin secretion differs from the I1 subclass and suggest that moxonidine inhibits insulin secretion by a mechanism unrelated to imidazoline I1 receptor agonism.     

Proteolytic release of membrane proteins: studies on a membrane-protein-solubilizing activity in CHO cells

Diverse membrane proteins are solubilized by a specific proteolytic cleavage in the stalk sequence adjacent to the membrane anchor, with release of the extracellular domain. Examples are the amyloid precursor protein, membrane-bound growth factors and angiotensin-converting enzyme (ACE). The identities and characteristics of the responsible proteases remain elusive. We have studied this process in Chinese hamster ovary (CHO) cells stably expressing wild-type ACE (WT-ACE) or juxtamembrane (stalk) deletion or chimaera mutants. Determination of the C termini (i.e. the cleavage sites) of released, soluble wild-type and mutant ACE by MALDI-TOF mass spectrometry indicated that the membrane-protein-solubilizing protease (MPSP) in CHO cells is not constrained by a particular cleavage site motif or by a specific distance from the membrane, but instead may position itself with respect to the putative proximal, folded extracellular domain adjacent to the stalk. Nevertheless, kinetic analyses of release rates indicated that a minimum distance from the membrane must be preserved. Interestingly, soluble full-length (anchor-plus) WT-ACE incubated with fractions of, or intact, CHO cells was not cleaved. In all cases, release was stimulated by a media change or by the addition of phorbol ester, with rate enhancements of 5- and 50-fold, respectively, for WT-ACE. The phorbol ester effect was abolished by staurosporine, a protein kinase C (PKC) inhibitor. We propose that the CHO cell MPSP that solubilizes ACE: (1) only cleaves proteins embedded in a membrane; (2) requires an accessible stalk and cleaves at a minimum distance from both the membrane and proximal extracellular domain; (3) positions itself primarily with respect to the proximal extracellular domain and (4) is regulated in part by a PKC-dependent mechanism.