The insect homologue of the amyloid precursor protein interacts with the heterotrimeric G protein Go alpha in an identified population of migratory neurons.

The amyloid precursor protein (APP) is the source of Abeta fragments implicated in the formation of senile plaques in Alzheimer's disease (AD). APP-related proteins are also expressed at high levels in the embryonic nervous system and may serve a variety of developmental functions, including the regulation of neuronal migration. To investigate this issue, we have cloned an orthologue of APP (msAPPL) from the moth, Manduca sexta, a preparation that permits in vivo manipulations of an identified set of migratory neurons (EP cells) within the developing enteric nervous system. Previously, we found that EP cell migration is regulated by the heterotrimeric G protein Goalpha: when activated by unknown receptors, Goalpha induces the onset of Ca2+ spiking in these neurons, which in turn down-regulates neuronal motility. We have now shown that msAPPL is first expressed by the EP cells shortly before the onset of migration and that this protein undergoes a sequence of trafficking, processing, and glycosylation events that correspond to discrete phases of neuronal migration and differentiation. We also show that msAPPL interacts with Goalpha in the EP cells, suggesting that msAPPL may serve as a novel G-protein-coupled receptor capable of modulating specific aspects of migration via Goalpha-dependent signal transduction.

Calcium deficiency in fluoride-treated osteoporotic patients despite calcium supplementation.

To test the hypothesis that the osteogenic response to fluoride can increase the skeletal requirement for calcium, resulting in a general state of calcium deficiency and secondary hyperparathyroidism, we assessed calcium deficiency, spinal bone density, by quantitative computed tomography, and serum PTH in three groups of osteoporotic subjects. Two of the three groups had been treated with fluoride and calcium (at least 1500 mg/day) for 32 +/- 19 months. Group 1 consisted of 16 fluoride-treated subjects who had shown rapid increases in spinal bone density (+ 3.8 +/- 2.6 mg/cm2 month), group II consisted of 10 fluoride-treated subjects who had shown decreases or only slow increases in spinal bone density (-0.05 +/- 0.6 mg/cm3 month), and group III consisted of 10 age-matched untreated osteoporotic controls. Calcium deficiency was assessed by measurement of calcium retention after calcium infusion. The results of our studies showed that 1) 94% of the subjects in Group I were calcium deficient compared with only 30% in groups II and III (P < 0.01 for each); 2) the subjects in group I retained more calcium (79%) than the subjects in group II (60%, P < 0.001) or the subjects in group III (64%, P < 0.005); 3) calcium retention was proportional to serum PTH (r = 0.37, n = 36, P < 0.03); and 4) calcium retention was proportional to the (previous) fluoride-dependent increase in quantitative computed tomography spinal bone density (in groups I and II, r = 0.48, n = 26, P < 0.02). To test the hypothesis that the calcium deficiency and the secondary hyperparathyroidism that were associated with the positive response to fluoride would respond to concomitant calcitriol treatment, a subgroup of 7 calcium-deficient subjects were selected from group I and treated with calcitriol (plus fluoride and calcium) for an average of 7 months. The calcitriol therapy reduced the calcium deficit in all 7 subjects, decreasing calcium retention from 80% to 62% (P < 0.02), and decreasing PTH from 50 to 28 pg/mL (P < 0.02). Together, these data indicate that fluoride-treated osteoporotic subjects may develop calcium deficiency in proportion to the effect of fluoride to increase bone formation, and this calcium deficit is responsive to calcitriol therapy.

The increase in spinal bone density that occurs in response to fluoride therapy for osteoporosis is not maintained after the therapy is discontinued.

In 44 osteoporotic subjects who had been treated with fluoride for 37 +/- 16 months, the fluoride was discontinued because they had shown fluoride-dependent increases in trabecular spinal bone densities from low initial levels (below the fracture threshold) to values that were equivalent to normal peak bone densities in the spines of young adults. During the subsequent period, after discontinuation of the fluoride therapy (i.e. 19 +/- 9 months), spinal bone density decreased in 73% of the subjects (i.e. 32 of 44, p < 0.03), at a rate that was comparable to the rate of the previous gain that had occurred during the treatment with fluoride (i.e. -3.23 +/- 2.39 mg/cm3 per month, compared with + 3.91 +/- 1.96 mg/cm3 per month in this, subgroup of patients, p < 0.001). Although 9 of the 44 subjects showed continuing increases in spinal bone density after discontinuation of the fluoride therapy, spinal bone density decreased in the entire group of 44 at an average rate of -1.02 +/- 4.72 mg/cm3 per month (p < 0.001, compared with the rate of the previous gain during the treatment with fluoride; i.e. +3.83 +/- 1.82 mg/cm3 per month). Surprisingly, our data showed that the rate of decrease in spinal bone density during the post-fluoride period was not affected by concurrent (undesigned) treatment with calcium, calcium plus estrogen, or calcium plus calcitriol. The cessation of fluoride therapy was also associated with a decrease in serum alkaline phosphatase activity (i.e. a decrease from the elevated levels that were observed during the period of fluoride therapy, back to the original, pre-treatment levels; p < 0.001), and that the rate of spinal bone loss after cessation of fluoride could be correlated with the prior rate of increase in serum alkaline phosphatase activity that had occurred during the treatment with fluoride (n = 44, r = 0.312, p = 0.039). Together, the observations from this retrospective analysis of data obtained from our clinical subjects suggest that fluoride-treated osteoporotic subjects who have exhibited increases in trabecular spinal bone density are at risk for bone loss after discontinuation of the fluoride therapy.

Lack of a high prevalence of the BB vitamin D receptor genotype in severely osteoporotic women.

Studies of twins strongly suggest that more than 50% of the peak spinal bone density is determined by genetics. It was reported recently that this genetic effect is primarily determined by vitamin D receptor (VDR) alleles; specifically, a VDR genotype termed BB has been highly associated with low peak bone density. Homozygotes for the second VDR allele, bb, are associated with high peak bone density. If peak bone density is an important determinant of osteoporosis and if the VDR genotype is an important determinant of peak bone density, then patients with severe osteoporosis should have a high prevalence of the BB VDR genotype compared with that of control subjects. To test this hypothesis, we used Southern blot analysis to determine the VDR genotype of 41 Caucasian patients (72 +/- 14 yr) with severe osteoporosis (27 women with spinal bone densities below 50 mg/cm3 as determined by quantitative computed tomography; 14 women with spinal bone densities below 0.75 g/cm2 as determined by dual energy x-ray absorptiometry) and 23 Caucasian control subjects (68 +/- 7 yr) without osteoporosis (quantitative computed tomography values at or above the fracture threshold of 100 mg/cm3). Only 6 of the 41 individuals in the group with severe osteoporosis had the BB genotype, whereas 16 had the bb genotype. In the control group comprising 23 individuals, 7 had the BB genotype and only 6 had the bb genotype. We conclude that the BB VDR genotype is not a good predictor of risk for developing severe osteoporosis in our population.

Spinal fractures during fluoride therapy for osteoporosis: relationship to spinal bone density.

Recent studies report that fluoride therapy for osteoporosis increases spinal bone density without improving vertebral fracture rate, challenging the notion that restoration of bone mass improves bone fragility. To further evaluate this issue, the relationship between spinal bone density and vertebral fracture rate was examined in a large number of fluoride-treated, osteoporotic patients. A retrospective assessment was made of clinical data collected from our observations of 389 osteoporotics treated with fluoride 30 +/- 8 mg/day (mean +/- SD) (equivalent to 66 +/- 17 mg NaF/day) and calcium 1500 mg/day for 28 +/- 18 months. Fracture rate and bone density were assessed in the same region of the spine (i.e., T12 through L4) using quantitative computed tomography (QCT). Spinal bone density increased with time on fluoride, but the relationship was hyperbolic (r = 0.99, p less than 0.0001; asymptote = 167 mg/cc on double-reciprocal plot), suggesting a plateau in the response. The spinal fracture rate decreased as a function of time on therapy (r = -0.83, p less than 0.01), and was inversely related to spinal bone density during fluoride therapy (r = 0.70, p less than 0.001 on arithmetic plot; r = -0.79, p less than 0.001 on semi-log plot). The subgroup of patients who responded to treatment with a significant increase in spinal bone density had a 48% reduction in spinal fracture rate compared with non-responders (p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Fluoride therapy for osteoporosis: a review of dose response, duration of treatment, and skeletal sites of action.

Osteoporosis is a disease characterized by a reduction in bone density which predisposes to fracture after even minimal trauma. Fluoride, because it has consistently been shown to stimulate bone formation and increase trabecular bone density, has been widely studied for the treatment of osteoporosis. The article focuses on the dose response, duration of treatment, and skeletal sites of action of fluoride; we also include comments on the effect of fluoride on vertebral and appendicular fracture rates. The skeletal response to fluoride doses, ranging from 15 to 43 mg elemental fluoride per day, included a linear increase in spinal bone density at an average rate of 1.25 +/- 0.91 mg/cm3 per month. The rate of increase in spinal bone density was related to the dose of fluoride (r = 0.34, P less than 0.03). Spinal bone density had increased above the fracture threshold in 44% of patients treated with fluoride for 32 +/- 10 months. The time required to achieve this goal was, however, influenced by the pretreatment spinal bone density and interpatient variation in response to fluoride treatment. Patients whose spinal bone density remained below the fracture threshold had lower pretreatment bone densities and/or slower rates of increase in spinal bone density (P less than 0.001). The osteogenic effect of fluoride was not limited to the spine. After 2 years of fluoride therapy, we found bone density in the femoral condyle (measured by QCT) to have increased by 13 +/- 2.6 mg/cm3 (n = 38, P less than 0.001); bone density in the hip (measured by DPA) was increased by 0.0261 +/- 0.015 g/cm2 (n = 55, P less than 0.025).(ABSTRACT TRUNCATED AT 250 WORDS)

Fluoride therapy for osteoporosis promotes a progressive increase in spinal bone density.

Since osteoporosis is a disease of diminished bone density, and since osteoporotic fractures occur most commonly in the spine, the ideal therapeutic agent for osteoporosis is one which can increase spinal bone density and thereby reduce the risk for vertebral fractures. In the current study we sought to examine the effect of fluoride therapy on spinal bone density utilizing quantitative computed tomography to measure changes in vertebral trabecular bone density during treatment with fluoride. A group of 61 postmenopausal osteoporotic females, aged 70 +/- 9 years, were treated with 34 +/- 7 mg elemental fluoride/day (equivalent to 75 +/- 15 mg NaF/day) and 1500 mg calcium/day for 19 +/- 6 months. Spinal bone density was increased within the first 6 months of fluoride therapy by 42% or 10 +/- 13 mg/cm3 (p less than 0.001) and continued to increased throughout 2 years of observation. The skeletal response to fluoride therapy was also associated with an early increase in serum alkaline phosphatase activity (p less than 0.001), which was related to the increase in spinal bone density (r = .58, p less than 0.001). Large interpatient variation was observed in the spinal bone response to fluoride therapy, which was not explained by variations in the pretreatment spinal bone density (r = .04), age of the patient (r = .15), or dose of fluoride (r = .16). Results from these studies demonstrate (1) the therapeutic value of fluoride to increase trabecular bone density linearly for 2 years in the osteoporotic spine and (2) the clinical value of measuring spinal bone density and/or serum alkaline phosphatase activity as indices of the skeletal response to fluoride.

Fluoride pharmacokinetics in good and poor responders to fluoride therapy.

In this study, the relationship between fluoride pharmacokinetics and the response in spinal bone density to fluoride treatment was studied in 14 patients with primary osteoporosis treated with fluoride for at least 1 year. Serum concentrations and urinary excretion of fluoride were determined after ingestion of 10 mg fluoride as monofluorophosphate. The pharmacokinetic parameters were calculated according to a linear one-compartment open model. The fasting serum fluoride level was 8.8 +/- 0.98 mumol/liter. The peak serum fluoride level was 20.5 +/- 1.4 mumol/liter and was reached within 2 h after ingestion of fluoride. When the patients were divided into good and poor responders, based on whether they did or did not exhibit a change in spinal bone density of 13 mg/cc per year or more, we found that good responders had decreased renal fluoride clearance (-62 +/- 13%, p less than .02), increased maximum change in serum fluoride (+38 +/- 18%, p less than .01), increased extrarenal clearance (+62 +/- 57%, p less than .05) and increased change in serum alkaline phosphatase (ALP) (+241 +/- 169%, p less than 0.02) compared with poor responders. Our data suggest that one factor accounting for a good response is a relatively high serum level of fluoride. However, although the maximum change in serum fluoride was greater in good responders compared with poor responders, variations in fluoride levels could not explain all of the variation in spinal bone density. Therefore, we propose that in addition to differences in serum fluoride, other factors are also responsible for the good response.

Efficacy of long-term fluoride and calcium therapy in correcting the deficit of spinal bone density in osteoporosis.

Long-term fluoride therapy for osteoporosis has been shown to increase the thickness of vertebral trabeculae as seen on spinal radiographs. To determine if this qualitative finding represents a measurable increase in spinal bone density, quantitative computed tomography was utilized to measure trabecular vertebral body density (TVBD) in the lumbar spine of 18 female osteoporotic patients, all of whom had been treated with sodium fluoride, 77 +/- 13 mg/day (mean +/- SD), and calcium, 1000 mg/day, for 57 +/- 24 months. TVBD in these fluoride treated osteoporotic patients (132 +/- 82 mg/cm3) was found to be significantly greater than mean TVBD for an age-matched group of untreated female osteoporotic patients (51 +/- 21 mg/cm3, n = 89, p less than 0.001). The value for TVBD in the long-term fluoride treated osteoporotics was not only similar to previously published values for TVBD (104 +/- 30 mg cm3) in normal females of similar age, but was also above the calculated TVBD "fracture threshold" of 100 mg/cm3 for females. Only one of the 18 fluoride treated osteoporotics continued to have spinal fractures during therapy, accounting for 4 fractures per 87.2 patient years of observation, a value which is significantly lower than the published incidence of 76 fractures per 91 patient years for untreated osteoporotic patients (p less than 0.001). Together, these findings demonstrate that long-term fluoride and calcium therapy for osteoporosis increases TVBD in the majority of patients within a reasonable time frame and significantly reduces the risk for spinal fractures.(ABSTRACT TRUNCATED AT 250 WORDS)

Fluoride therapy for osteoporosis: characterization of the skeletal response by serial measurements of serum alkaline phosphatase activity.

Optimum use of fluoride therapy for osteoporosis requires a sensitive and convenient index of the skeletal response to fluoride. Since previous studies had shown that serum alkaline phosphatase activity (SALP) was increased in response to fluoride therapy, we examined serial measurements of SALP in 53 osteoporotics treated with 66 to 110 mg of sodium fluoride (NaF) for 12 to 91 months. SALP was increased in 87% of the subjects during therapy with fluoride. The increase in SALP was thought to reflect the osteogenic action of fluoride based on the findings that SALP correlated with both trabecular bone area (r = .81, P less than .001) and osteoid length (r = .67, P less than .01) in iliac crest biopsies, predicted increased bone density on spinal radiographs in response to fluoride therapy with an 87% accuracy, and predicted decreased back pain in response to fluoride with a 91% accuracy. In addition, the SALP response to fluoride was seen earlier than other therapeutic responses as indicated by the findings that the tau 1/2 for the SALP response (ie, time for 1/2 of the patients to show a significant response) was significantly less (1.2 +/- 0.3 yr) than that for the pain response (1.6 +/- 0.3 yr, P less than .05) or that for the radiographic response (3.7 +/- 0.5 yr, P less than .001). Although most patients responded to fluoride with an increase in SALP, evaluation of the kinetics of the SALP response to fluoride revealed marked interpatient variation.(ABSTRACT TRUNCATED AT 250 WORDS)

Skeletal scintigraphic changes in osteoporosis treated with sodium fluoride: concise communication.

An appendicular skeletal response to sodium fluoride (NaF) was detected by total skeletal scintigrams. Twelve postmenopausal osteoporotic women were treated with NaF (88 mg/day) and calcium (1500 mg/day). Total skeletal scintigrams were obtained before and during treatment. Within 4 to 21 mo (mean: 8.3), all 12 patients showed new areas of increased uptake corresponding to metaphyseal regions and short bones of the appendicular skeleton. The number of peripheral bones involved in each subject ranged from four to 12. The most frequently involved sites (11 of 12 patients) were the right distal femur and proximal tibia. Nine patients showed an increase in serum alkaline phosphatase activity, which was attributed to an increase in the skeletal isoenzyme. Seven of 12 patients developed bone pain in one or more of the regions of increased uptake. This study establishes that the skeletal scintigram is a sensitive index of the peripheral skeletal response to NaF.