Measuring vitamin D levels: surrogates are insufficient.

AIMS: With the increasing evidence of adverse consequences because of low vitamin D levels on health demand for vitamin D, screening is increasing. The objective of the study was to assess whether parathyroid hormone (PTH) levels/bone profile is sufficient to identify patients with vitamin D insufficiency or deficiency, or whether vitamin D should be measured directly. METHODOLOGY: A total of 1560 serum specimens, with requests for 25-hydroxyvitamin D (25-OH vitamin D), calcium, phosphate, alkaline phosphatase (ALP), creatinine and PTH on the same sample were analysed at Salford Royal Hospital from November 2010 to November 2012. RESULTS: The prevalence of total vitamin D insufficiency or deficiency (defined as total 25-OH vitamin D < 50 nmol/l) was 62.9% (981/1560) overall, with males having higher proportions (67.2 vs. 59.3 per cent; χ(2) = 8.78, p = 0.003). There was no overall trend in mean serum adjusted calcium across categories of 25-OH vitamin D status but mean serum phosphate was significantly lower (F = 6.53, p < 0.0001) in patients with a 25-OH vitamin D level < 50 nmol/l. However in patients with vitamin D deficiency, a significant proportion had PTH, calcium, phosphate and alkaline phosphatase levels within the laboratory normal range. Even at a 25-OH vitamin D < 10 nmol/l, 71.6% had a normal PTH, 89.8% had normal serum calcium levels, 84.9% had normal phosphate levels and 81.6% had normal serum ALP. CONCLUSIONS: Therefore, despite the costs associated with the measurement of vitamin D, our findings show that no surrogate is adequate for screening for vitamin D deficiency.

Reducing Risks for a Dissatisfied Patient in Facial Cosmetic Surgery.

Patient selection in aesthetic surgery is the ultimate inexact science, a mixture of surgical judgment, expertise, ego, gut feelings, personality interactions, and a spin of the roulette wheel. Using our procedural skills as well as our interpersonal skills will enhance our professional satisfaction and improve the quality of life for others as we try to reduce the likelihood of dissatisfaction, both for our patients and ourselves.

Avoiding and correcting complications in facial plastic surgery: patient selection.

Far from being simply a cursory step, the initial consultation between the patient and plastic surgeon, when patient selection usually occurs, can be complicated. Proper patient selection is the key first step toward a successful outcome from a proposed procedure. Not only must the physical and health status of the patient be understood, but the surgeon quickly must determine whether the emotional and psychological attributes and expectations of a particular patient are up to the rigors of the particular surgery being discussed. The concepts of expectations and satisfaction, personality types, and the difficult patient will be discussed. Yes, predicting the future is part of the surgeon's valued armamentaria. But even experienced and astute physicians cannot always make perfect decisions.

Identification of a novel cell type-specific intronic enhancer of macrophage migration inhibitory factor (MIF) and its regulation by mithramycin.

The aim of this study was to determine the genetic regulation of macrophage migration inhibitory factor (MIF). DNase I hypersensitivity was used to identify potential hypersensitive sites (HS) across the MIF gene locus. Reporter gene assays were performed in different human cell lines with constructs containing the native or mutated HS element. Following phylogenetic and transcription factor binding profiling, electrophoretic mobility shift assay (EMSA) and RNA interference were performed and the effects of incubation with mithramycin, an antibiotic that binds GC boxes, were also studied. An HS centred on the first intron of MIF was identified. The HS acted as an enhancer in human T lymphoblasts (CEMC7A), human embryonic kidney cells (HEK293T) and human monocytic cells (THP-1), but not in a fibroblast-like synoviocyte (FLS) cell line (SW982) or cultured FLS derived from rheumatoid arthritis (RA) patients. Two cis-elements within the first intron were found to be responsible for the enhancer activity. Mutation of the consensus Sp1 GC box on each cis-element abrogated enhancer activity and EMSA indicated Sp1 binding to one of the cis-elements contained in the intron. SiRNA knock-down of Sp1 alone or Sp1 and Sp3 together was incomplete and did not alter the enhancer activity. Mithramycin inhibited expression of MIF in CEMC7A cells. This effect was specific to the intronic enhancer and was not seen on the MIF promoter. These results identify a novel, cell type-specific enhancer of MIF. The enhancer appears to be driven by Sp1 or related Sp family members and is highly sensitive to inhibition via mithramycin.

Interferon-inducible factor 16 is a novel modulator of glucocorticoid action.

Previously, we used cDNA expression profiling to identify genes associated with glucocorticoid (Gc) sensitivity. We now identify which of these directly influence Gc action. Interferon-inducible protein 16 (IFI16), bone morphogenetic protein receptor type II (BMPRII), and regulator of G-protein signaling 14 (RGS14) increased Gc transactivation, whereas sialyltransferase 4B (SIAT4B) had a negative effect. Amyloid beta (A4) precursor-protein binding, family B, member 1 (APBB1/Fe65) and neural cell expressed developmentally down-regulated 9 (NEDD9) were without effect. Only IFI16 potentiated Gc repression of NF-kappaB. In addition, IFI16 affected basal expression, and Gc induction of endogenous target genes. IFI16 did not affect glucocorticoid receptor (GR) expression, ligand-dependent repression of GR expression, or the ligand-dependent induction of GR phosphorylation on Ser-211 or Ser-203. Coimmunoprecipitation revealed an interaction, suggesting that IFI16 modulation of GR function is mediated by protein crosstalk. Transfection analysis with GR mutants showed that the ligand-binding domain of GR binds IFI16 and is the target domain for IFI16 regulation. Analysis of human lung sections identified colocalization of GR and IFI16, suggesting a physiologically relevant interaction. We demonstrate that IFI16 is a novel modulator of GR function and show the importance of analyzing variation in Gc sensitivity in humans, using appropriate technology, to drive discovery.

Overcoming barriers to EHR adoption.

Implementing an electronic health record (EHR) system that links medical facilities in three states taught PeaceHealth these valuable lessons: Establish clinical standards. Talk to clinicians early on. Look for an EHR with clinical decision support capabilities.

Functional evaluation of TNFAIP3 (A20) in rheumatoid arthritis.

OBJECTIVES: To determine the protein expression of TNFAIP3 in synovium and to show the capability of 6q23 intergenic SNPs, associated with rheumatoid arthritis (RA) susceptibility, to influence TNFAIP3 gene transcription. METHODS: Immunohistochemistry for TNFAIP3, NF-kB p65 and phosphorylated NF-kB p65 protein expression was performed in 6 RA knee joint synovium samples compared to 9 osteoarthritis (OA) samples. Luciferase reporter gene assays were used to examine the regulatory ability of RA associated SNP variants on TNFAIP3 promoter activity. Sense and antisense constructs were prepared for rs6920220 alleles, together with each of the 4 SNPs in r2=1 with it (rs6933404, rs2327832, rs6927172 and rs17264332), coupled to the TNFAIP3 promoter. Transient transfections were performed in a human T lymphoblastoid (CEMC7A) cell line. Bioinformatic software was utilised to prioritise SNPs for further investigation. Electrophoretic mobility shift assays (EMSA), using CEMC7A nuclear extracts, were conducted for the rs6927172 SNP alleles. RESULTS: TNFAIP3 protein expression was seen in the synovium samples and differential TNFAIP3 protein expression between RA vs. OA synoviocytes observed. Within RA synoviocytes TNFAIP3 expression is predominately cytoplasmic, whereas in OA its expression is strongly nuclear and cytoplasmic. For 3 of the 5 SNPs investigated (rs6920220, rs6933404, rs6927172) evidence of repressor activity of TNFAIP3 transcription was seen and EMSA data showed evidence of differential transcription factor binding to rs6927172 alleles. CONCLUSIONS: This is the first observation of TNFAIP3 protein expression in RA and OA synovium. In vitro analysis of 6q23 intergenic SNPs supports the possibility of the functional regulation of TNFAIP3.

It's a jungle out there: survival guide for the facial plastic surgeon.

Many different people present to the surgeon for possible elective procedures, but not all of them are suitable patients. Learning to identify possible problematic patients before surgery is a valuable survival strategy in the profession of plastic surgery. In this article, an "animal model" is used whereby patient and physician "species" are presented in a lighthearted tongue-in-cheek manner but also in a practical sense.

Essays on patient management and elective surgery.

This article presents a series of essays from the author with thoughts and opinions pertaining to the practice of facial plastic surgery and how surgeons deal with patients who anticipate often impossible procedures.

Roundtable: discussion of "the difficult patient".

Eight renowned surgeons responded to questions centering on "difficult patients" in facial plastic surgery. Questions ranged from, "How do you manage a postoperative patient who looks 'OK,' if not great, to you but complains about the result?" to "What 'pearl of advice' would you offer a novice surgeon on how to best avoid difficult situations with their patients?" The surgeons taking part in the discussion, from different practices in different parts of the country, provided a lively discussion based on their years of experience.

Macrophage migration inhibitory factor (MIF) and oligoarticular juvenile idiopathic arthritis (o-JIA): association of MIF promoter polymorphisms with response to intra-articular glucocorticoids.

OBJECTIVES: To address the clinical relevance of macrophage migration inhibitory factor (MIF) promoter polymorphisms in oligoarticular juvenile idiopathic arthritis (o-JIA) by evaluating their associations with serum and SF MIF levels, with response to intra-articular glucocorticoid injections and with outcome of the disease. METHODS: Seventy-five Caucasian patients with o-JIA were studied. Alleles of the -794 CATT variable number of tandem repeats (VNTR) and of the -173 G/C single nucleotide polymorphism (SNP) were identified by capillary electrophoresis following fluorescently labelled PCR and by allelic discrimination assay, respectively. MIF levels were measured by ELISA. The association of MIF promoter polymorphisms with polyarticular extension, Childhood Health Assessment Questionnaire (CHAQ) score at the last follow-up visit and occurrence of chronic anterior uveitis was evaluated only in patients with a follow up > 5 years. RESULTS: Neither of the MIF promoter polymorphisms was associated with serum MIF levels, nor with the long-term outcome of o-JIA. The -173 G/C SNP was significantly associated with both SF MIF levels and duration of response to intra-articular glucocorticoid injection. Carriers of a MIF -173 C allele were 4 times more likely to relapse within 3 months. No association was found between the different MIF CATT alleles and both SF MIF levels and duration of response to intra-articular glucocorticoids. CONCLUSION: Our study shows the clinical relevance of the MIF -173 G/C SNP in o-JIA and suggests that the -173 C allele may represent a predictor of poor response to intra-articular glucocorticoid treatment.

Glucocorticoids suppress macrophage migration inhibitory factor (MIF) expression in a cell-type-specific manner.

MIF is a potent proinflammatory cytokine involved in inflammatory arthritis. Glucocorticoids (GC) have been reported to induce secretion of MIF in rodent cells, and as MIF counteracts the anti-inflammatory effects of GC, this has implications for human inflammatory disease. Transient transfection studies showed that the MIF promoter was repressed by dexamethasone (Dex) (10 nM) in CEM C7A cells, with up to 50% suppression by 100 nM. However, there was no regulation of the promoter by GC in A549 cells. We also found that subnanomolar concentrations of Dex suppressed MIF secretion, measured by ELISA, by 80% in both human T lymphoblasts (CEM C7A) and human lung epithelial cells (A549). Endogenous MIF mRNA was also repressed by GC in CEM C7A cells, measured both by Northern blot and quantitative RT-PCR assays, but there was no such regulation in A549 cells. This suggests that GC affects translation rather than transcription of MIF in A549 cells. These results contradict earlier results with the rat cell line RAW 264.7. Therefore, we analysed MIF secretion from RAW 264.7 cells but found no GC effect on secretion. Understanding how GC regulates MIF in a cell-type-dependent manner may give insights into GC-refractory human inflammatory diseases.

Interleukin-10 (IL10) promoter polymorphisms and multiple sclerosis.

Interleukin-10 (IL10) is an anti-inflammatory cytokine which may modulate disease expression in multiple sclerosis (MS). Three dimorphic polymorphisms within the IL10 promoter region at positions - 1082, -819 and -519 have previously been identified. The - 1082*A allele has been associated with low and the - 1082*G allele with high in vitro IL10 production. We have genotyped 185 Caucasian MS patients and 211 ethnically matched controls for each of these three dimorphisms. MS patients were stratified for severity of disease outcome. No associations were found for any IL10 promoter polymorphisms when the MS cases were compared with controls or with disease outcome with regards to disability. IL10 polymorphism does not appear to be associated with MS or to influence disease progression.

Macrophage migration inhibitory factor: molecular, cellular and genetic aspects of a key neuroendocrine molecule.

The immunological and neuroendocrine properties of macrophage migration inhibitory factor (MIF) are diverse. In this article we review the known cellular, molecular and genetic properties of MIF that place it as a key regulatory cytokine, acting within both the innate and adaptive immune responses. The unexpected and paradoxical induction of MIF secretion by low concentrations of glucocorticoids is explored. The role of MIF as a locally acting modulator of glucocorticoid sensitivity within foci of inflammation is also discussed. MIF has no homology with any other pro-inflammatory cytokine and until recently lacked a recognised transmembrane receptor. MIF has also been shown to be directly taken up into target cells and to interact with intracellular signalling molecules, including the Jun activation domain-binding protein Jab-1.Comprehensive analysis of the MIF gene has identified important functional polymorphisms and a series of genetic studies has revealed both association and linkage of MIF with inflammatory diseases. Altered MIF regulation may therefore be pivotal to acquiring chronic inflammation following an innate immune response.

HLA-DRB1 and disease outcome in multiple sclerosis.

The association between susceptibility to multiple sclerosis (MS) and the class II MHC allele HLA-DRB1*15 is well established although a possible relationship between this allele and outcome in MS is less clear. HLA-DRB1 typing was performed on 375 unrelated white patients with clinically definite MS and on 367 healthy controls. Putative associations of the gene with outcome were examined by dividing patients into two groups: those with an EDSS of 0-5.5 (mild/moderate disease) and those with an EDSS of 6-10 (severe disease). In order to minimise the effects of disease variability patients with a disease duration of at least 10 years or 15 years were examined. As subsidiary HLA-DRB1*03 and HLA-DRB1*04 associations have been previously reported, the effect of these alleles was also examined. As expected, HLA-DRB1*15 was found more frequently in patients than in controls (P < 0.000001). HLA-DRB1*15 positive patients had a significantly earlier age at onset than HLA-DRB1*15 negative patients. No significant associations were noted between HLA-DRB1*15 and outcome in the total patient group or in patients with a disease duration of 10 years or longer. In patients with a disease duration of at least 15 years HLA-DRB1*15 negative status was associated with a worse prognosis, although this did not remain significant after correction for multiple testing. It is thus likely that the contribution of HLA in MS is primarily towards onset and initial triggering mechanisms rather than influencing disease progression, chronicity and severity.

TAP2D is associated with HLA-B44 and DR4 and may contribute to rheumatoid arthritis and Felty's syndrome susceptibility.

OBJECTIVES: TAP2 transporter gene polymorphisms have been ascertained in patients with rheumatoid arthritis (RA) and Felty's syndrome (FS) to determine whether particular alleles of this gene are disease associated. METHODS: TAP2 dimorphisms at amino acid positions 379, 565 and 665 were detected using ARMS-PCR in 89 RA patients, 24 FS patients and 64 control subjects. TAP 2 alleles were assigned from these results. RESULTS: The frequency of one particular allele, TAP2D, was increased in both RA (OR 2.6, 95% CI 1.2 - 5.8) and FS (OR 3.9, 95% CI 1.4 - 10.7). When individual amino acid polymorphisms were compared between patients and controls, isoleucine at position 379 (present in TAP2D and TAP2C) was significantly increased, indicating that this dimorphism itself may be associated with RA (OR 5.0, 95% CI 2.4 - 10.2) and FS (OR 5.0, 95% CI 1.91 - 3.2). DISCUSSION: The presence of TAP2D was greatly increased in HLA-B44/DR4 positive RA (83%) and FS (67%) patients. These frequencies were appreciably higher than in the HLA-B44/DR4 controls (11%), suggesting that linkage disequilibrium alone may not explain the increase in TAP2D frequency in patients and that this allele may represent an additional risk factor in these conditions.

Network analysis: a new approach to study endocrine disorders.

Systems biology is the study of the interactions that occur between the components of individual cells - including genes, proteins, transcription factors, small molecules, and metabolites, and their relationships to complex physiological and pathological processes. The application of systems biology to medicine promises rapid advances in both our understanding of disease and the development of novel treatment options. Network biology has emerged as the primary tool for studying systems biology as it utilises the mathematical analysis of the relationships between connected objects in a biological system and allows the integration of varied 'omic' datasets (including genomics, metabolomics, proteomics, etc.). Analysis of network biology generates interactome models to infer and assess function; to understand mechanisms, and to prioritise candidates for further investigation. This review provides an overview of network methods used to support this research and an insight into current applications of network analysis applied to endocrinology. A wide spectrum of endocrine disorders are included ranging from congenital hyperinsulinism in infancy, through childhood developmental and growth disorders, to the development of metabolic diseases in early and late adulthood, such as obesity and obesity-related pathologies. In addition to providing a deeper understanding of diseases processes, network biology is also central to the development of personalised treatment strategies which will integrate pharmacogenomics with systems biology of the individual.