Did the UK's public health shielding policy protect the clinically extremely vulnerable during the COVID-19 pandemic in Wales? Results of EVITE Immunity, a linked data retrospective study.

INTRODUCTION: The UK shielding policy intended to protect people at the highest risk of harm from COVID-19 infection. We aimed to describe intervention effects in Wales at 1 year. METHODS: Retrospective comparison of linked demographic and clinical data for cohorts comprising people identified for shielding from 23 March to 21 May 2020; and the rest of the population. Health records were extracted with event dates between 23 March 2020 and 22 March 2021 for the comparator cohort and from the date of inclusion until 1 year later for the shielded cohort. RESULTS: The shielded cohort included 117,415 people, with 3,086,385 in the comparator cohort. The largest clinical categories in the shielded cohort were severe respiratory condition (35.5%), immunosuppressive therapy (25.9%) and cancer (18.6%). People in the shielded cohort were more likely to be female, aged ≥50 years, living in relatively deprived areas, care home residents and frail. The proportion of people tested for COVID-19 was higher in the shielded cohort (odds ratio [OR] 1.616; 95% confidence interval [CI] 1.597-1.637), with lower positivity rate incident rate ratios 0.716 (95% CI 0.697-0.736). The known infection rate was higher in the shielded cohort (5.9% vs 5.7%). People in the shielded cohort were more likely to die (OR 3.683; 95% CI: 3.583-3.786), have a critical care admission (OR 3.339; 95% CI: 3.111-3.583), hospital emergency admission (OR 2.883; 95% CI: 2.837-2.930), emergency department attendance (OR 1.893; 95% CI: 1.867-1.919) and common mental disorder (OR 1.762; 95% CI: 1.735-1.789). CONCLUSION: Deaths and healthcare utilisation were higher amongst shielded people than the general population, as would be expected in the sicker population. Differences in testing rates, deprivation and pre-existing health are potential confounders; however, lack of clear impact on infection rates raises questions about the success of shielding and indicates that further research is required to fully evaluate this national policy intervention.

The effect of nitrogen containing bisphosphonates, zoledronate and alendronate, on the production of pro-angiogenic factors by osteoblastic cells.

Bisphosphonates (BPs) have been shown to influence angiogenesis. This may contribute to BP-associated side-effects such as osteonecrosis of the jaw (ONJ) or atypical femoral fractures (AFF). The effect of BPs on the production of angiogenic factors by osteoblasts is unclear. The aims were to investigate the effect of (1) alendronate on circulating angiogenic factors; vascular endothelial growth factor (VEGF) and angiopoietin-1 (ANG-1) in vivo and (2) zoledronate and alendronate on the production of VEGF and ANG-1 by osteoblasts in vitro. We studied 18 post-menopausal women with T score⩽-2 randomized to calcium/vitamin D only (control arm, n=8) or calcium/vitamin D and alendronate 70mg weekly (treatment arm, n=10). Circulating concentrations of VEGF and ANG-1 were measured at baseline, 3, 6 and 12months. Two human osteoblastic cell lines (MG-63 and HCC1) and a murine osteocytic cell line (MLO-Y4) were treated with zoledronate or alendronate at concentrations of 10(-12)-10(-6)M. VEGF and ANG-1 were measured in the cell culture supernatant. We observed a trend towards a decline in VEGF and ANG-1 at 6 and 12months following treatment with alendronate (p=0.08). Production of VEGF and ANG-1 by the MG-63 and HCC1 cells decreased significantly by 34-39% (p<0.01) following treatment with zoledronate (10(-9)-10(-6)M). Treatment of the MG-63 cells with alendronate (10(-7) and 10(-6)) led to a smaller decrease (25-28%) in VEGF (p<0.05). Zoledronate (10(-10)-10(-)(6)M) suppressed the production of ANG-1 by MG-63 cells with a decrease of 43-49% (p<0.01). Co-treatment with calcitriol (10(-8)M) partially reversed this zoledronate-induced inhibition. BPs suppress osteoblastic production of angiogenic factors. This may explain, in part, the pathogenesis of the BP-associated side-effects.

Preclinical assessment of a new magnetic resonance-based technique for determining bone quality by characterization of trabecular microarchitecture.

The utility of HR-CT to study longitudinal changes in bone microarchitecture is limited by subject radiation exposure. Although MR is not subject to this limitation, it is limited both by patient movement that occurs during prolonged scanning at distal sites, and by the signal-to-noise ratio that is achievable for high-resolution images in a reasonable scan time at proximal sites. Recently, a novel MR-based technique, fine structure analysis (FSA) (Chase et al. Localised one-dimensional magnetic resonance spatial frequency spectroscopy. PCT/US2012/068284 2012, James and Chase Magnetic field gradient structure characteristic assessment using one-dimensional (1D) spatial frequency distribution analysis. 7932720 B2, 2011) has been developed which provides both high-resolution and fast scan times, but which generates at a designated set of spatial positions (voxels) a one-dimensional signal of spatial frequencies. Appendix 1 provides a brief introduction to FSA. This article describes an initial exploration of FSA for the rapid, non-invasive characterization of trabecular microarchitecture in a preclinical setting. For L4 vertebrae of sham and ovariectomized (OVX) rats, we compared FSA-generated metrics with those from CT datasets and from CT-derived histomorphometry parameters, trabecular number (Tb.N), bone volume density (BV/TV), trabecular thickness (Tb.Th) and trabecular separation (Tb.Sp). OVX caused a reduction of the higher frequency structures that correspond to a denser trabecular lattice, while increasing the preponderance of lower frequency structures, which correspond to a more open lattice. As one example measure, the centroid of the FSA spectrum (which we refer to as fSAcB) showed strong correlation in the same region with CT-derived histomorphometry values: Tb.Sp: r -0.63, p < 0.001; Tb.N: r 0.71, p < 0.001; BV/TV: r 0.64, p < 0.001, Tb.Th: r 0.44, p < 0.05. Furthermore, we found a 17.5% reduction in fSAcB in OVX rats (p < 0.0001). In a longitudinal study, FSA showed that the age-related increase in higher frequency structures was abolished in OVX rats, being replaced with a 78-194% increase in lower frequency structures (2.4-2.8 objects/mm range), indicating a more sparse trabecular lattice (p < 0.05). The MR-based fine structure analysis enables high-resolution, radiation-free, rapid quantification of bone structures in one dimension (the specific point and direction being chosen by the clinician) of the spine.

Biomimetic cilia arrays generate simultaneous pumping and mixing regimes.

Living systems employ cilia to control and to sense the flow of fluids for many purposes, such as pumping, locomotion, feeding, and tissue morphogenesis. Beyond their use in biology, functional arrays of artificial cilia have been envisaged as a potential biomimetic strategy for inducing fluid flow and mixing in lab-on-a-chip devices. Here we report on fluid transport produced by magnetically actuated arrays of biomimetic cilia whose size approaches that of their biological counterparts, a scale at which advection and diffusion compete to determine mass transport. Our biomimetic cilia recreate the beat shape of embryonic nodal cilia, simultaneously generating two sharply segregated regimes of fluid flow: Above the cilia tips their motion causes directed, long-range fluid transport, whereas below the tips we show that the cilia beat generates an enhanced diffusivity capable of producing increased mixing rates. These two distinct types of flow occur simultaneously and are separated in space by less than 5 microm, approximately 20% of the biomimetic cilium length. While this suggests that our system may have applications as a versatile microfluidics device, we also focus on the biological implications of our findings. Our statistical analysis of particle transport identifying an enhanced diffusion regime provides novel evidence for the existence of mixing in ciliated systems, and we demonstrate that the directed transport regime is Poiseuille-Couette flow, the first analytical model consistent with biological measurements of fluid flow in the embryonic node.

Contrasting effects of A1 and A2b adenosine receptors on adipogenesis.

BACKGROUND: Adenosine mediates its actions through four G protein-coupled receptors, A1, A2a, A2b and A3. The A1 receptor (A1R) is dominant in adipocytes where it mediates many actions that include inhibition of lipolysis, stimulation of leptin secretion and protection against obesity-related insulin resistance. OBJECTIVE: The objective of this study is to investigate whether induced expression of A1Rs stimulates adipogenesis, or whether A1R expression is a consequence of cells having an adipocyte phenotype. METHODOLOGY: Human A1R and A2b receptors (A2bRs) were stably transfected into a murine osteoblast precursor cell line, 7F2. Adipogenesis was determined by lipid accumulation and expression of adipocyte and osteoblast marker molecules. Adenosine receptor expression and activation of associated signal molecules were also evaluated as 7F2 cells were induced to differentiate to adipocytes. RESULTS: 7F2 cells transfected with the A1R showed increased adipocyte marker mRNA expression; lipoprotein lipase and glycerol-3-phosphate dehydrogenase were both upregulated, whereas the osteoblast marker alkaline phosphatase (ALP) was downregulated. When cultured in adipocyte differentiating media, such cells also showed increased adipogenesis as judged by lipid accumulation. Conversely, A2bR transfection stimulated osteocalcin and ALP expression, and in addition, adipogenesis was inhibited in the presence of adipocyte differentiation media. Adipogenic differentiation of naive 7F2 cells also resulted in increased expression of the A1R and reduced or modified expression of the A2a and A2bR. The loss of A2 receptors after adipogenic differentiation was accompanied by a loss of cyclic adenosine monophosphate and ERK1/2 signalling. CONCLUSION: These data show that expression of A1Rs induced adipocyte differentiation, whereas A2bR expression inhibited adipogenesis and stimulated an osteoblastic phenotype. These data suggest that targeting A1 and A2bR could be considered in the management of obesity and diabetes. Targeting adenosine signal pathways may be useful in treatment strategies for diseases in which there is an imbalance between osteoblasts and adipocytes.

A Streptococcus pneumoniae infection model in larvae of the wax moth Galleria mellonella.

The bacterium Streptococcus pneumoniae is a leading human opportunistic pathogen. The limitations of the current vaccine have led to increased recognition of the need to understand bacterial behaviour and competitive dynamics using in vivo models of infection. Here, we investigate the potential application of the larvae of the wax moth Galleria mellonella as an informative infection model. Larvae were challenged with a range of doses of S. pneumoniae isolates differing in known virulence factors to determine the LD(50) values. Infection dynamics were determined by obtaining bacterial counts from larvae over a time course. Differences in virulence between serotypes could be distinguished in this host. Infection with strains differing in known virulence factors demonstrated predicted differences in virulence. Acapsulate and pneumolysin-negative strains were less virulent than their respective wild types. A large reduction in virulence was seen in strains lacking cell wall D-alanylation. The mortality of G. mellonella larvae is attributable to bacterial growth within larvae, while surviving larvae are able to clear infections by reducing bacterial numbers. These data demonstrate that G. mellonella larvae represent an in vivo infection model with applications for investigating aspects of bacterial-host interactions such as the role of antimicrobial peptide activity and resistance.

Novel genetic context of multiple bla OXA-58 genes in Acinetobacter genospecies 3.

OBJECTIVES: The detection in Acinetobacter genospecies 3 isolates of OXA-type carbapenemases, resulting in reduced susceptibility to carbapenem antibiotics, is increasingly reported. We identified an Acinetobacter genospecies 3 isolate carrying the gene for OXA-58 and aimed to resolve the genetic environment surrounding the bla(OXA-58) gene. METHODS: Species identification was confirmed by 16S-23S rRNA restriction analysis. MICs of imipenem, meropenem and ertapenem were determined, and the isolate was screened by PCR for bla(OXA-23-like), bla(OXA-40-like), bla(OXA-51-like) and bla(OXA-58-like) genes. The sequence surrounding bla(OXA-58) was determined through amplification by inverse PCR and genome walking followed by sequencing. Genetic localization was investigated by Southern blotting. RESULTS: Isolate A164 was confirmed as belonging to Acinetobacter genospecies 3 and had reduced susceptibility to the carbapenems. The isolate was found to encode two bla(OXA-58) genes that may have been duplicated by the insertion sequence ISAba125, two copies of which were inserted into ISAba3 elements. The bla(OXA-58) genes appear to be plasmid borne. CONCLUSIONS: This is the first report of beta-lactamase duplication in Acinetobacter genospecies 3 and of gene duplication mediated by ISAba125.

Fitness correlates with the extent of cheating in a bacterium.

There is growing awareness of the importance of cooperative behaviours in microbial communities. Empirical support for this insight comes from experiments using mutant strains, termed 'cheats', which exploit the cooperative behaviour of wild-type strains. However, little detailed work has gone into characterising the competitive dynamics of cooperative and cheating strains. We test three specific predictions about the fitness consequences of cheating to different extents by examining the production of the iron-scavenging siderophore molecule, pyoverdin, in the bacterium Pseudomonas aeruginosa. We create a collection of mutants that differ in the amount of pyoverdin that they produce (from 1% to 96% of the production of paired wild types) and demonstrate that these production levels correlate with both gene activity and the ability to bind iron. Across these mutants, we found that (1) when grown in a mixed culture with a cooperative wild-type strain, the relative fitness of a mutant is negatively correlated with the amount of pyoverdin that it produces; (2) the absolute and relative fitness of the wild-type strain in the mixed culture is positively correlated with the amount of pyoverdin that the mutant produces; and (3) when grown in a monoculture, the absolute fitness of the mutant is positively correlated with the amount of pyoverdin that it produces. Overall, we demonstrate that cooperative pyoverdin production is exploitable and illustrate how variation in a social behaviour determines fitness differently, depending on the social environment.

OXA-51-like beta-lactamases and their association with particular epidemic lineages of Acinetobacter baumannii.

Sixty diverse clinical Acinetobacter baumannii isolates of worldwide origin were assigned to sequence groups, based on a multiplex PCR for the ompA, csuE and bla(OXA-51-like) genes. The majority (77%) of isolates belonged to sequence groups 1 and 2 (SG1 and SG2), with sequence group 3 (SG3) and non-grouped isolates accounting for the remainder. The isolates were not closely related according to pulsed-field gel electrophoresis (PFGE), and the majority were sensitive to imipenem and meropenem. The construction of a linkage map of OXA-51-like beta-lactamase sequence relationships revealed two closely related clusters of enzymes, one focused around OXA-66 and the other around OXA-69. Isolates belonging to SG1 encoded an enzyme from the OXA-66 cluster, while those belonging to SG2 encoded an enzyme from the OXA-69 cluster. All SG3 isolates encoded OXA-71, which does not form part of a close enzyme grouping. Major multinational lineages accounted for a significant proportion of A. baumannii clinical isolates, and the evolution of the OXA-51-like enzymes appears to be an ongoing process.

Magnetic microkayaks: propulsion of microrods precessing near a surface by kilohertz frequency, rotating magnetic fields.

Surface-swimming nano- and micromotors hold significant potential for on-chip mixing, flow generation, sample manipulation, and microrobotics. Here we describe rotating microrods magnetized nearly orthogonally to their long axes. When actuated near a solid surface, these microrods demonstrate precessing motion, with rods describing a double cone similar to the motion of a kayaker's paddle. The precessing motion induces translation. At 1 kHz, these "microkayaks" move at translational velocities of ≈14 µm s-1 and generate advective flows up to 10 µm s-1.

Effects of an air pollution personal alert system on health service usage in a high-risk general population: a quasi-experimental study using linked data.

BACKGROUND: There is no evidence to date on whether an intervention alerting people to high levels of pollution is effective in reducing health service utilisation. We evaluated alert accuracy and the effect of a targeted personal air pollution alert system, airAware, on emergency hospital admissions, emergency department attendances, general practitioner contacts and prescribed medications. METHODS: Quasi-experimental study describing accuracy of alerts compared with pollution triggers; and comparing relative changes in healthcare utilisation in the intervention group to those who did not sign-up. Participants were people diagnosed with asthma, chronic obstructive pulmonary disease (COPD) or coronary heart disease, resident in an industrial area of south Wales and registered patients at 1 of 4 general practices. Longitudinal anonymised record linked data were modelled for participants and non-participants, adjusting for differences between groups. RESULTS: During the 2-year intervention period alerts were correctly issued on 208 of 248 occasions; sensitivity was 83.9% (95% CI 78.8% to 87.9%) and specificity 99.5% (95% CI 99.3% to 99.6%). The intervention was associated with a 4-fold increase in admissions for respiratory conditions (incidence rate ratio (IRR) 3.97; 95% CI 1.59 to 9.93) and a near doubling of emergency department attendance (IRR=1.89; 95% CI 1.34 to 2.68). CONCLUSIONS: The intervention was associated with increased emergency admissions for respiratory conditions. While findings may be context specific, evidence from this evaluation questions the benefits of implementing near real-time personal pollution alert systems for high-risk individuals.

Alpha-inhibin gene expression occurs in the ovine adrenal cortex, and is regulated by adrenocorticotropin.

Inhibin is a glycoprotein hormone composed of two nonidentical subunits. It is produced by the ovary and testis and plays a vital role in gonadal function by inhibiting the secretion of FSH. More recently, additional activities associated with inhibin peptides have been identified. Inhibin heterodimers (alpha-beta) are reported to act directly on ovarian granulosa cells and inhibit estrogen production induced by FSH. Furthermore, homodimers of beta-inhibin subunits stimulate the secretion of FSH, an activity that is directly opposite to that of inhibin. Each of these inhibin-related activities are concerned with the hypothalamic-pituitary-gonadal axis. We have investigated further the complexity of inhibin activity by determining whether inhibin genes are expressed in nongonadal tissue. RNA hybridization experiments demonstrate that the alpha-inhibin gene is expressed in the sheep adrenal cortex and hybridization histochemistry shows that this gene is expressed in each of the functional zones within the cortex. Dot blot analysis showed that the level of alpha mRNA within the adrenal is influenced by ACTH, one of the major regulators of adrenal cortex function. These observations imply that there are inhibin-related peptides not directly associated with the gonads. beta-inhibin gene expression was not clearly detected in the adrenal and we conclude that if expression occurs then it does so at extremely low levels.

Complete androgen insensitivity due to deletion of exon C of the androgen receptor gene highlights the functional importance of the second zinc finger of the androgen receptor in vivo.

Androgen-dependent gene transcription is mediated by the androgen receptor (AR) through interaction of its central zinc finger region with specific DNA sequences on target genes. Failure of this receptor-mediated gene transcription results in end organ resistance to androgens-the androgen insensitivity syndromes. In a pair of siblings with complete androgen insensitivity who had supranormal levels of androgen binding in genital skin fibroblasts, polymerase chain reaction and Southern blot analysis of the androgen receptor gene confirmed by polymerase chain reaction and sequence analysis of AR cDNA, revealed an in-frame deletion of exon C encoding the second zinc finger of the receptor. The mutant receptor in cultured genital skin fibroblasts had normal androgen binding affinity and was localized in the nucleus but had markedly reduced DNA-binding affinity. When recreated in vitro and tested in a cotransfection assay system the mutant receptor failed to activate transcription of an androgen-responsive reporter gene. This naturally occurring mutation highlights the functional dependence of the AR upon its second zinc finger in vivo and explains the complete insensitivity to androgen manifest by the affected individuals despite increased androgen binding. The elevated AR levels in the subjects' genital skin fibroblasts further suggests a possible role for the second zinc finger in autoregulation of receptor levels in vivo.

'Diaphragmlike' stricture and ulcer of the colon during diclofenac treatment.

Diclofenac sodium is a widely used enteric-coated nonsteroidal anti-inflammatory drug. We describe a woman with Hemoccult-positive stools and iron deficiency anemia who developed both a colonic ulcer and a "diaphragm-like" colonic stricture while taking enteric-coated diclofenac. These lesions were evident on colonoscopy but not on barium studies. Biopsy specimens of the ulcer and stricture revealed particulate matter that was indistinguishable from diclofenac pill fragments by electron microscopy. Discontinuation of diclofenac therapy resulted in resolution of anemia and Hemoccult-positive stools. We conclude that (1) enteric-coated diclofenac is associated with both colonic ulcers and diaphragm-like colonic strictures; (2) the pathophysiologic mechanism for the development of both ulcers and strictures may involve a direct action of diclofenac within these lesions; (3) colonoscopy may be superior to barium studies in evaluating patients receiving diclofenac who have iron deficiency anemia and/or Hemoccult-positive stools.

Benign tertiary syphilis and HIV infection.

Benign tertiary (gummatous) syphilis has virtually disappeared [1]. Two homosexual men with gummata have presented to genitourinary medicine clinics in west London in recent months. They both had antibodies to HIV. It is possible that otherwise adequately treated syphilis has progressed to the tertiary stage as a consequence of HIV infection and modification of the immune response.

Morphological determinants of femoral strength in growth hormone-deficient transgenic growth-retarded (Tgr) rats.

UNLABELLED: The extent to which childhood GHD affects adult fracture risk is unclear. We measured femoral strength in adult transgenic growth-retarded rats as a model of GHD. Long-term, moderate GHD was accompanied by endocrine and morphometric changes consistent with a significant reduction in femoral strength. INTRODUCTION: Childhood growth hormone deficiency (GHD) is associated with osteopenia, but little is known about its effects on subsequent adult bone strength and fracture risk. MATERIALS AND METHODS: We have therefore measured femoral strength (failure load measured by three-point bending) in a new model of moderate GHD, the transgenic growth-retarded (Tgr) rat at 15, 22-23, and 52 weeks of age, and have quantified potential morphological and endocrine determinants of bone strength. RESULTS: Skeletal growth retardation in Tgr rats was accompanied by a sustained reduction in the anterior-posterior diameter of the femoral cortex, whereas mid-diaphyseal cortical wall thicknesses were largely unaltered. Total femoral strength was significantly impaired in Tgr rats (p < 0.01), and this impairment was more pronounced in males than females. Compromised bone strength in Tgr rats could not be accounted for by the reduction in mechanical load (body weight) and was not caused by impairment of the material properties of the calcified tissue (ultimate tensile stress), despite marked reductions in femoral mineral density (areal bone mineral density; p < 0.001). Microcomputerized tomographical analysis revealed significant modification of the architecture of trabecular bone in Tgr rats, with reductions in the number and thickness of trabeculae (p < 0.05) and in the degree of anisotropy (p < 0.01). The marked reduction in plasma insulin-like growth factor-1 in Tgr rats was accompanied by the development of high circulating leptin levels (p < 0.01). CONCLUSION: These results show that the changes in endocrinology and bone morphology associated with long-term moderate GHD in Tgr rats are accompanied by changes consistent with a significant reduction in the threshold for femoral fracture.

Structural and biochemical effects of essential fatty acid deficiency on peripheral nerve.

The effect of postweaning essential fatty acid (EFA) deficiency on the peripheral nerve was studied in groups of rats. At 325 days, the characteristic biochemical changes of EFA deficiency were present in isolated peripheral myelin, although to a lesser degree than reported in non-neural tissues. There was no significant difference between control and deficient groups in number or size distributions of myelinated fibers (MFs) in muscle and sensory nerves, in the incidence of teased fiber abnormalities, in rates of axonal transport of dopamine-beta-hydroxylase and acetylcholinesterase, or in conduction velocity and compound action potentials of peripheral nerve in vivo or in vitro. Four weeks after a standard sciatic crush injury, the median MF diameter in regenerated peroneal nerves was significantly smaller in EFA-deficient rats than in control rats, but this difference was no longer significant at 18 weeks. At 18 weeks, EFA-deficient and control regenerated nerves showed similar myelin periodicity and relationship of axonal area to number of myelin lamellae. We conclude that acquired EFA deficiency in the rat leads to biochemically abnormal peripheral myelin, but that this state is unaccompanied by clinical, physiological, or morphological evidence of neuropathy.

Complete androgen insensitivity syndrome characterized by increased concentration of a normal androgen receptor in genital skin fibroblasts.

Two siblings with the classical phenotype of complete androgen insensitivity syndrome (CAIS) and increased total cellular androgen receptor concentrations in genital skin fibroblasts (GSF) are described. Testosterone biosynthesis was normal, and there was no evidence of 5 alpha-reductase deficiency. Specific binding of [3H]dihydrotestosterone ([3H]DHT) in GSF was 7 SD above the mean value in normal fibroblast strains [maximum binding, 775 +/- 185 X 10(-18) mol/micrograms DNA (mean +/- SD)]. Binding at 40 C was stable, and the androgen-receptor complex dissociated at a normal rate (t1/2, 85 min). The androgen-receptor complex from GSF cytosol sedimented at 5-6S on sucrose density gradients in the presence of sodium molybdate. In a whole cell binding assay, the percentage of [3H]DHT that bound to a crude nuclear pellet was 60%. Preincubation of GSF with 2 nM [3H]DHT for 20 h before the standard 1-h whole cell binding assay produced a further augmentation in elevated total cellular androgen receptor concentrations. A new variant of CAIS is described which is characterized by an increased concentration of androgen receptors that appear to be quantitatively and qualitatively normal. Augmentation of the receptor by androgen suggests that the gene coding for the androgen receptor is intact and does not account for the androgen insensitivity.

Effects of chemotherapeutic agents on the function of primary human osteoblast-like cells derived from children.

Studies in children treated with chemotherapy suggest that chemotherapeutic agents have deleterious effects on bone metabolism. We therefore evaluated the in vitro effects of clinically relevant concentrations of chemotherapeutic agents on the synthesis of type I collagen, alkaline phosphatase (AP) activity, and mineralization by primary human osteoblast-like (HOB) cells derived from children. Because serum 1,25-dihydroxyvitamin D(3) concentrations may be reduced during treatment with chemotherapy, the effect of chemotherapeutic agents on HOB cells cultured in the presence or absence of 1,25-dihydroxyvitamin D(3) was also evaluated. Type I collagen synthesis was reduced by all agents (P < 0.01) other than methotrexate, whereas the relative AP activity was increased (P < 0.01) by all agents. The relative number of cells staining intensely for AP after culture with agents increased (P < 0.05), and AP mRNA expression was increased (P < 0.01) with vincristine. 1,25-Dihydroxyvitamin D(3) ameliorated (P < 0.01) the depletion of HOB cell numbers by chemotherapeutic agents. Furthermore, vincristine and daunorubicin inhibited 1,25-dihydroxyvitamin D(3)-mediated AP activity (P < 0.01). We conclude that chemotherapeutic agents can adversely affect HOB cell function, and we speculate that this observation may account, in part, for the osteopenia observed during and after treatment of children with chemotherapy.