Ramasamyiyer Swaminathan

Genetic Contribution to Bone Metabolism, Calcium Excretion, and Vitamin D and Parathyroid Hormone Regulation

A classical twin study was performed to assess the relative contribution of genetic and environmental factors to bone metabolism, calcium homeostasis, and the hormones regulating them. It was examined further whether the genetic effect is menopause dependent. The subjects were 2136 adult twins (98.3% female): 384 monozygotic (MZ) and 684 dizygotic (DZ) twin pairs. The intraclass correlations were calculated, and maximum likelihood model fitting was used to estimate genetic and environmental variance components. The intraclass correlations for all of the variables assessed were higher in MZ twin pairs. The heritabilities (95% CIs) obtained from model fitting for hormones regulating bone metabolism and calcium homeostasis were parathyroid hormone (PTH), 60% (54–65%); 25‐hydroxyvitamin D [25(OH)D]; 43% (28–57%), 1,25‐hydroxyvitamin D [1,25(OH)], 65% (53–74%); and vitamin D binding protein 62% (56–66%). The heritabilities (95% CIs) for markers of bone formation also were assessed; bone‐specific alkaline phosphatase (BSAP), 74% (67–80%), and osteocalcin, 29% (14–44%); marker of bone resorption deoxypyridinoline (DPD), 58% (52–64%); and measure of calcium homeostasis 24 h urine calcium, creatinine (Cr), 52% (41–61%). The magnitude of genetic influence differed with menopause for most variables. This study provides evidence for the importance of genetic factors in determining bone resorption and formation, calcium excretion, and the hormones regulating these processes. It shows for the first time a clear genetic effect on bone resorption in premenopausal women and the regulation of PTH, vitamin D metabolism, and calcium excretion. The genes controlling bone hormones and markers are likely to be useful therapeutic and diagnostic targets.

Self-monitoring in Type 2 diabetes mellitus: a meta-analysis

SUMMARY

Influence of raised plasma osmolality on clinical outcome after acute stroke

Background and Purpose Abnormal physiological parameters after acute stroke may induce early neurological deterioration. Studies of the effect of dehydration on stroke outcome are limited. We examined the association of raised plasma osmolality on stroke outcome at 3 months and the change of plasma osmolality with hydration during the first week after stroke. Methods Acute stroke patients had their plasma osmolality measured at admission and at days 1, 3, and 7. Maximum plasma osmolality and the area under curve (AUC) were also calculated during the first week. Patients were stratified according to how they were hydrated: orally, intravenously, or both. Outcome included survival at 3 months after stroke. Logistic regression was performed to examine the association between raised plasma osmolality (>296 mOsm/kg) and survival, adjusting for stroke severity. Linear regression was performed to examine the pattern of plasma osmolality across hydration groups. Results One hundred sixty-seven patients were included. Mean admission (300 mOsm/kg, SD 11.4), maximum (308.1 mOsm/kg, SD 17.1), and AUC (298.3 mOsm/kg, SD 11.7) plasma osmolality were significantly higher in those who died compared with survivors (293.1 mOsm/kg [SD 8.2], 297.7 mOsm/kg [SD 8.7], and 291.7 mOsm/kg [SD 8.1], respectively;P <0.0001). Admission plasma osmolality >296 mOsm/kg was significantly associated with mortality (OR 2.4, 95% CI 1.0 to 5.9). In patients hydrated intravenously, there was no significant fall in plasma osmolality compared with patients hydrated orally (P =0.68). Conclusions Raised plasma osmolality on admission is associated with stroke mortality, after correcting for case mix. Correction of dehydration after stroke requires a more systematic approach. Trials are required to determine whether correcting dehydration after stroke improves outcome.

Biochemical markers of bone turnover

BACKGROUND Osteoporosis in many countries has reached epidemic proportions. This has stimulated the development of biochemical markers to assist in the assessment of osteoporotic risk and in monitoring the efficacy of treatment. Biochemical markers of bone turnover are products released from osteoblasts and osteoclasts or collagen breakdown products. MARKERS Markers of bone formation include bone-specific alkaline phosphatase (BAP), osteocalcin (OC) and procollagen peptides. All of these can be measured easily by immunoassay techniques. Of these markers, OC has been extensively studied. However, OC undergoes in vitro degradation, thus, assay results are variable. BAP, on the other hand, is much more stable and shows less within-person biological variation. Bone resorption markers include tartrate-resistant acid phosphatase (TRAP) and collagen breakdown products, such as pyridinium cross-links, galactosyl hydroxylysine and cross-linked telopeptides, such as CTx and NTx. Of these, deoxypyridinium (DPD) has been extensively studied. DPD shows diurnal variation and the within-individual biological variation is large. Of the newer assays, NTx appear to show large differences at menopause. CONCLUSIONS Thus, serum BAP and DPD or NTx are the current choice of bone markers.

Circulating Nucleic Acids in Plasma and Serum : Recent Developments

Abstract:  Nucleic acids (DNA and RNA) have been detected in plasma, serum, urine, and other body fluids from healthy subjects as well as in patients. The ability to detect and quantitate specific DNA and RNA sequences has opened up the possibility of diagnosis and monitoring of diseases. With the recent developments in the field of circulating nucleic acids the application in the diagnostic field has increased. The recent discovery of epigenetic changes in placental/fetal DNA and the detection of fetal/placental‐specific RNAs have made it possible to use this technology in all pregnancies irrespective of the gender of the fetus. With the application of mass spectrometry and other techniques to this field, it is now possible to detect very small amounts of specific DNA in the presence of excess of other nonspecific nucleic acids (e.g., detection of mutations in fetal DNA in the presence of excess of maternal DNA). Circulating nucleic acids have now been shown to be useful in other conditions, such as diabetes mellitus, trauma, stroke, and myocardial infarction. In oncology, detection and monitoring of tumors is now possible by the detection of tumor‐derived nucleic acids. In spite of these advances questions regarding the origin and biologic significance of circulating nucleic acids remain to be answered. Furthermore preanalytical and analytical aspects of this field remain to be standardized.

Circulating Nucleic Acids in Plasma and Serum

DNA and RNA fractions have been isolated from the whole blood, serum, plasma, the surface of blood cells, and urine of both healthy individuals and patients. The ability to isolate, quantify, and analyze these molecules has led to the identification of specific nucleic acid fragments related to particular disorders such as diabetes, cancer, myocardial infarction, and stroke, threby permitting their early diagnosis. Currently, a number of methods for isolating the nucleic acids are employed and although a start has been made to compare the efficiencies of these methods, there is still a way to go before there are precise protocols for nucleic acid extraction. The older chemical methods of extraction still outperform some of the available kits. Some progress is being made to determine the origin of the circulating nucleic acids, although there are still many questions to be answered, including whether the source is through the spontaneous release of newly synthesized nucleic acid or whether it just derived from necrotic and apoptotic cells. In addition, it can be demonstrated that the nucleic acids can enter cells and exhibit a biological activity in the recipient cells. Hence, the question remains: Are the circulating nucleic acids freely entering tissues and cells from the blood and inducing changes in those tissues and cells? Further work is needed to elucidate these areas, and the various protocols must be standardized if the new methodology is to be widely and accurately applied in the diagnosis of disease and the monitoring of therapy. This chapter summarizes the work reported in this volume.

Reactivation of tuberculosis and vitamin D deficiency: the contribution of diet and exposure to sunlight

Background: As well as its role in the regulation of calcium metabolism, vitamin D is an immunoregulatory hormone. Epidemiological evidence also suggests a link between vitamin D deficiency and tuberculosis (TB). A study was undertaken to examine serum vitamin D concentrations before treatment in patients with active TB and their contacts from the same ethnic and social background and to investigate the relative contributions of diet and sunlight exposure. Methods: Serum vitamin D concentrations were measured before treatment in 178 patients with active TB and 130 healthy contacts. The prevalence of vitamin D deficiency and its relation to skin colour, month of estimation and TB diagnosis were determined. 35 patients and 35 frequency-matched contacts completed dietary and sun exposure questionnaires to determine the relative contribution of these to serum vitamin D concentrations. Results: There was a statistically significant difference in serum vitamin D concentrations between patients and contacts (20.1 vs 30.8 nmol/l, 95% CI 7.1 to 14.3; p<0.001) and significantly more patients had severely deficient concentrations (<21 nmol/l) than controls (114/178 (64%) vs 40/130 (31%), p<0.001). There was no association between serum concentrations of vitamin D and skin pigmentation. The healthy contacts showed a predictable seasonal pattern, rising to peak concentrations in the summer months, but this response was absent in patients with TB. Dietary intake was the same in both patients with TB and contacts matched for age, sex and skin colour, but patients with TB displayed a stronger correlation between serum vitamin D concentrations and dietary intake (r = 0.42, p = 0.016) than controls (r = 0.13, p>0.1). There was no difference in sunlight exposure between the groups. Conclusions: Patients with active TB have lower serum vitamin D concentrations than contacts from similar ethnic and social backgrounds and with comparable dietary intake and sun exposure, and do not show the expected seasonal variation. These observations indicate that other factors are contributing to vitamin D deficiency in patients with TB and suggest abnormal handling of this vitamin.

A Randomized Controlled Trial of Vitamin D Supplementation on Preventing Postmenopausal Bone Loss and Modifying Bone Metabolism Using Identical Twin Pairs

Vitamin D supplementation, when given with calcium, has been shown to increase bone mineral density (BMD) and reduce the incidence of hip fracture in elderly subjects. Despite its widespread use, the benefits of vitamin D supplementation in younger women and as a single agent are less clear. We performed a randomized co‐twin, placebo‐controlled, double‐blind trial over 2 years to measure the effect of vitamin D3 supplementation on bone density and bone metabolism in young postmenopausal women. Seventy‐nine monozygotic (MZ) twin pairs (mean age, 58.7 years; range, 47–70 years) were recruited. For each twin pair, one was randomized to 800 IU cholecalciferol/day for 2 years and the other was randomized to placebo. BMD was measured at the spine and hip and heel ultrasound at baseline, 12, 18, and 24 months. Samples were collected at 0, 3, and 6 months to measure serum calcium, 25‐hydroxyvitamin D [25(OH)D], parathyroid hormone (PTH), osteocalcin, and urinary deoxypyridinoline (DPD). In total, 64 pairs completed the study. No differences in baseline characteristics were seen between the groups. At 6 months, the treatment group had an increase in serum vitamin D [mean ± SEM intrapair difference, 14.1 ± 2.4 μg/liter (p < 0.001)]. There were no significant differences in other serum measurements or bone markers at 3 months or 6 months. At 24 months, no significant treatment effect was seen on BMD or calcaneal ultrasound change within pairs. Subanalysis of treatment response by vitamin D receptor (VDR) genotype revealed no significant difference in effect on BMD variables with treatment. On the basis of these results, vitamin D supplementation, on its own, cannot be recommended routinely as an osteoporosis prevention for healthy postmenopausal women with normal vitamin D levels under the age of 70 years.

Methotrexate inhibits the first committed step of purine biosynthesis in mitogen-stimulated human T-lymphocytes: a metabolic basis for efficacy in rheumatoid arthritis?

The immunosuppressive and anti-inflammatory effects of low-dose methotrexate (MTX) have been related directly to inhibition of folate-dependent enzymes by polyglutamated derivatives, or indirectly to adenosine release and/or apoptosis and clonal deletion of activated peripheral blood lymphocytes in S-phase. In this study of phytohaemagglutinin-stimulated primary human T-lymphocytes we show that MTX (20 nM to 20 microM) was cytostatic not cytotoxic, halting proliferation at G(1). This stasis of blastogenesis was associated with an inhibition of purine ribonucleotide synthesis but a stimulation of pyrimidine biosynthesis, the normal mitogen-induced expansion of ATP and GTP pools over 72 h being restricted to concentrations of unstimulated T-cells, whereas the increment in UTP pools exceeded that of controls. Decreased incorporation of H(14)CO(3) or [(14)C]glycine into purine ribonucleotides, with no radiolabel accumulation in any de novo synthetic intermediate but enhanced H(14)CO(3) incorporation into UTP, supported these MTX-related effects. Exaggerated [(14)C]hypoxanthine salvage (which normalized the purine and UTP pools) confirmed the increased availability of 5-phosphoribosyl-1-pyrophosphate (PP-ribose-P) as the molecular mechanism underlying these disparate changes. These results provide the first substantive evidence that the immunosuppressive effects of low-dose MTX in primary blasting human T-lymphocytes relate not to the inhibition of the two folate-dependent enzymes of purine biosynthesis but to inhibition of the first enzyme, amidophosphoribosyltransferase, thereby elevating PP-ribose-P and stimulating UTP synthesis. Varying cell types or incubation conditions employed by other workers, especially malignant/activated cells with high basal metabolic rates, might mask the effects noted in primary human T-lymphocytes. The findings imply the involvement of low-dose MTX in the inhibition of T-lymphocyte proliferation and proliferation-dependent processes in rheumatoid arthritis.

Vitamin D and bone mineral density

Bone mineral density (BMD) at the lumbar spine and the neck of femur and serum concentrations of 25-hydroxyvitamin D (25OHD), intact parathyroid hormone (PTH), alkaline phosphatase, calcium, albumin, creatinine and phosphate were measured in a group of 166 postmenopausal women (30–79 years) attending a bone clinic for bone density measurements. Four subjects with suspected primary hyperparathyroidism were excluded from analysis. BMD at the lumbar spine was correlated with body mass index (BMI) (r=0.278,p=0.0003), age (r=−0.194,p=0.0134) and serum 25OHD (r=0.188,p=0.0167). BMD at the neck of femur correlated with BMI (r=0.391,p<0.0001), age (r=−0.356,p<0.0001), PTH (r=−0.156,p=0.047) and serum 25OHD (r=0.231,p=0.0031). Stepwise multiple regression analysis showed that age, BMI and serum 25OHD contributed to the variation in BMD at lumbar spine. At the neck of femur, PTH was an additional contributor. We conclude that serum 25OHD makes a contribution to BMD a lumbar spine and neck of femur.