Rapid genomic changes by mineralotropic hormones and kinase SIK inhibition drive coordinated renal Cyp27b1 and Cyp24a1 expression via CREB modules.

Vitamin D metabolism centers on kidney regulation of Cyp27b1 by mineralotropic hormones, including induction by parathyroid hormone (PTH), suppression by fibroblast growth factor 23 (FGF23) and 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), and reciprocal regulations for Cyp24a1. This coordinated genomic regulation results in production of endocrine 1,25(OH)2D3, which, together with PTH and FGF23, controls mineral homeostasis. However, how these events are coordinated is unclear. Here, using in vivo chromatin immunoprecipitation sequencing in mouse kidney, we demonstrate that PTH activation rapidly induces increased recruitment of phosphorylated (p-133) CREB (pCREB) and its coactivators, CBP (CREB-binding protein) and CRTC2 (CREB-regulated transcription coactivator 2), to previously defined kidney-specific M1 and M21 enhancers near the Cyp27b1 gene. At distal enhancers of the Cyp24a1 gene, PTH suppression dismisses CBP with only minor changes in pCREB and CRTC2 occupancy, all of which correlate with decreased genomic activity and reduced transcripts. Treatment of mice with salt-inducible kinase inhibitors (YKL-05-099 and SK-124) yields rapid genomic recruitment of CRTC2 to Cyp27b1, limited interaction of CBP, and a transcriptional response for both Cyp27b1 and Cyp24a1 that mirrors the actions of PTH. Surprisingly, we find that 1,25(OH)2D3 suppression increases the occupancy of CRTC2 in the M1 enhancer, a novel observation for CRTC2 and 1,25(OH)2D3 action. Suppressive actions of 1,25(OH)2D3 and FGF23 at the Cyp27b1 gene are associated with reduced CBP recruitment at these CREB-module enhancers that disrupts full PTH induction. Our findings show that CRTC2 contributes to transcription of both Cyp27b1 and Cyp24a1, demonstrate salt-inducible kinase inhibition as a key modulator of vitamin D metabolism, and provide molecular insight into the coordinated mechanistic actions of PTH, FGF23, and 1,25(OH)2D3 in the kidney that regulate mineral homeostasis.

Adjusting COVID-19 Seroprevalence Survey Results to Account for Test Sensitivity and Specificity.

Population-based seroprevalence surveys can provide useful estimates of the number of individuals previously infected with serious acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and still susceptible, as well as contribute to better estimates of the case-fatality rate and other measures of coronavirus disease 2019 (COVID-19) severity. No serological test is 100% accurate, however, and the standard correction that epidemiologists use to adjust estimates relies on estimates of the test sensitivity and specificity often based on small validation studies. We have developed a fully Bayesian approach to adjust observed prevalence estimates for sensitivity and specificity. Application to a seroprevalence survey conducted in New York State in 2020 demonstrates that this approach results in more realistic-and narrower-credible intervals than the standard sensitivity analysis using confidence interval endpoints. In addition, the model permits incorporating data on the geographical distribution of reported case counts to create informative priors on the cumulative incidence to produce estimates and credible intervals for smaller geographic areas than often can be precisely estimated with seroprevalence surveys.

Exploring behavioural factors affecting nutritional supplement use among children in Honduras.

OBJECTIVE: This study explored social and behavioural factors associated with a home fortification of complementary foods program among families of undernourished children in 14 rural communities in Honduras. DESIGN: We collected and analysed survey data from a convenience sample of 196 households participating in a nutritional program using home fortification of complementary foods in 2017. The program supplied families with a soy-based atole powder fortified with micronutrients. A research team completed a face-to-face survey exploring social and behavioural factors associated with nutritional supplement use. Anthropometric measurements for participating children were abstracted from health clinic records of previous quarterly appointments. SETTING: The study took place in San Jose del Negrito, Honduras. PARTICIPANTS: Participants were parents or guardians of children enrolled in the nutrition program. RESULTS: Nearly half of participant families shared the nutritional supplement with other family members besides the index child, while 10 % reported using the supplement as a meal replacement for the child. Low education level of mothers was associated with improper use of the supplement (P = 0·005). Poorer families were more likely to share the supplement (P = 0·013). CONCLUSIONS: These results highlight the challenges of programs using home fortification of complementary foods in the context of food scarcity. Findings highlight the importance of increasing rural children's overall caloric intake, perhaps by increasing access to locally available protein sources. Results also suggest transitioning nutritional programs to family-based interventions to increase overall intended compliance to nutrition programming.

New Approaches to Assess Mechanisms of Action of Selective Vitamin D Analogues.

Recent studies of transcription have revealed an advanced set of overarching principles that govern vitamin D action on a genome-wide scale. These tenets of vitamin D transcription have emerged as a result of the application of now well-established techniques of chromatin immunoprecipitation coupled to next-generation DNA sequencing that have now been linked directly to CRISPR-Cas9 genomic editing in culture cells and in mouse tissues in vivo. Accordingly, these techniques have established that the vitamin D hormone modulates sets of cell-type specific genes via an initial action that involves rapid binding of the VDR-ligand complex to multiple enhancer elements at open chromatin sites that drive the expression of individual genes. Importantly, a sequential set of downstream events follows this initial binding that results in rapid histone acetylation at these sites, the recruitment of additional histone modifiers across the gene locus, and in many cases, the appearance of H3K36me3 and RNA polymerase II across gene bodies. The measured recruitment of these factors and/or activities and their presence at specific regions in the gene locus correlate with the emerging presence of cognate transcripts, thereby highlighting sequential molecular events that occur during activation of most genes both in vitro and in vivo. These features provide a novel approach to the study of vitamin D analogs and their actions in vivo and suggest that they can be used for synthetic compound evaluation and to select for novel tissue- and gene-specific features. This may be particularly useful for ligand activation of nuclear receptors given the targeting of these factors directly to genetic sites in the nucleus.

Targeted genomic deletions identify diverse enhancer functions and generate a kidney-specific, endocrine-deficient Cyp27b1 pseudo-null mouse.

Vitamin D3 is terminally bioactivated in the kidney to 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3) via cytochrome P450 family 27 subfamily B member 1 (CYP27B1), whose gene is regulated by parathyroid hormone (PTH), fibroblast growth factor 23 (FGF23), and 1,25(OH)2D3 Our recent genomic studies in the mouse have revealed a complex kidney-specific enhancer module within the introns of adjacent methyltransferase-like 1 (Mettl1) and Mettl21b that mediate basal and PTH-induced expression of Cyp27b1 and FGF23- and 1,25(OH)2D3-mediated repression. Gross deletion of these segments in mice has severe effects on Cyp27b1 regulation and skeletal phenotype but does not affect Cyp27b1 expression in nonrenal target cells (NRTCs). Here, we report a bimodal activity in the Mettl1 intronic enhancer with components responsible for PTH-mediated Cyp27b1 induction and 1,25(OH)2D3-mediated repression and additional activities, including FGF23 repression, within the Mettl21b enhancers. Deletion of both submodules eliminated basal Cyp27b1 expression and regulation in the kidney, leading to systemic and skeletal phenotypes similar to those of Cyp27b1-null mice. However, basal expression and lipopolysaccharide-induced regulation of Cyp27b1 in NRTCs was unperturbed. Importantly, dietary normalization of calcium, phosphate, PTH, and FGF23 rescued the skeletal phenotype of this mutant mouse, creating an ideal in vivo model to study nonrenal 1,25(OH)2D3 production in health and disease. Finally, we confirmed a conserved chromatin landscape in human kidney that is similar to that in mouse. These findings define a finely balanced homeostatic mechanism involving PTH and FGF23 together with protection from 1,25(OH)2D3 toxicity that is responsible for both adaptive vitamin D metabolism and mineral regulation.

A chromatin-based mechanism controls differential regulation of the cytochrome P450 gene Cyp24a1 in renal and non-renal tissues.

Cytochrome P450 family 27 subfamily B member 1 (CYP27B1) and CYP24A1 function to maintain physiological levels of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) in the kidney. Renal Cyp27b1 and Cyp24a1 expression levels are transcriptionally regulated in a highly reciprocal manner by parathyroid hormone (PTH), fibroblast growth factor 23 (FGF23), and 1,25(OH)2D3 In contrast, Cyp24a1 regulation in nonrenal target cells (NRTCs) is limited to induction by 1,25(OH)2D3 Herein, we used ChIP-Seq analyses of mouse tissues to identify regulatory regions within the Cyp24a1 gene locus. We found an extended region downstream of Cyp24a1 containing a cluster of sites, termed C24-DS1, binding PTH-sensitive cAMP-responsive element-binding protein (CREB) and a cluster termed C24-DS2 binding the vitamin D receptor (VDR). VDR-occupied sites were present in both the kidney and NRTCs, but pCREB sites were occupied only in the kidney. We deleted each segment in the mouse and observed that although the overt phenotypes of both cluster deletions were unremarkable, RNA analysis in the C24-DS1-deleted strain revealed a loss of basal renal Cyp24a1 expression, total resistance to FGF23 and PTH regulation, and secondary suppression of renal Cyp27b1; 1,25(OH)2D3 induction remained unaffected in all tissues. In contrast, loss of the VDR cluster in the C24-DS2-deleted strain did not affect 1,25(OH)2D3 induction of renal Cyp24a1 expression yet reduced but did not eliminate Cyp24a1 responses in NRTCs. We conclude that a chromatin-based mechanism differentially regulates Cyp24a1 in the kidney and NRTCs and is essential for the specific functions of Cyp24a1 in these two tissue types.

Antibody to Poly-N-acetyl glucosamine provides protection against intracellular pathogens: Mechanism of action and validation in horse foals challenged with Rhodococcus equi.

Immune correlates of protection against intracellular bacterial pathogens are largely thought to be cell-mediated, although a reasonable amount of data supports a role for antibody-mediated protection. To define a role for antibody-mediated immunity against an intracellular pathogen, Rhodococcus equi, that causes granulomatous pneumonia in horse foals, we devised and tested an experimental system relying solely on antibody-mediated protection against this host-specific etiologic agent. Immunity was induced by vaccinating pregnant mares 6 and 3 weeks prior to predicted parturition with a conjugate vaccine targeting the highly conserved microbial surface polysaccharide, poly-N-acetyl glucosamine (PNAG). We ascertained antibody was transferred to foals via colostrum, the only means for foals to acquire maternal antibody. Horses lack transplacental antibody transfer. Next, a randomized, controlled, blinded challenge was conducted by inoculating at ~4 weeks of age ~10(6) cfu of R. equi via intrabronchial challenge. Eleven of 12 (91%) foals born to immune mares did not develop clinical R. equi pneumonia, whereas 6 of 7 (86%) foals born to unvaccinated controls developed pneumonia (P = 0.0017). In a confirmatory passive immunization study, infusion of PNAG-hyperimmune plasma protected 100% of 5 foals against R. equi pneumonia whereas all 4 recipients of normal horse plasma developed clinical disease (P = 0.0079). Antibodies to PNAG mediated killing of extracellular and intracellular R. equi and other intracellular pathogens. Killing of intracellular organisms depended on antibody recognition of surface expression of PNAG on infected cells, along with complement deposition and PMN-assisted lysis of infected macrophages. Peripheral blood mononuclear cells from immune and protected foals released higher levels of interferon-γ in response to PNAG compared to controls, indicating vaccination also induced an antibody-dependent cellular release of this critical immune cytokine. Overall, antibody-mediated opsonic killing and interferon-γ release in response to PNAG may protect against diseases caused by intracellular bacterial pathogens.

Robotic Staged Bilateral Selective Postganglionic Sympathectomy for Upper-Extremity Hyperhidrosis.

PURPOSE: The prevalence of compensatory hyperhidrosis (CH) has been reported to be as high as 80% in patients following thoracic sympathectomy for upper-extremity hyperhidrosis. The CH rate is 7.2% with simultaneous bilateral robotic selective dorsal sympathectomy. We reviewed the results in patients who underwent staged bilateral robotic selective dorsal sympathectomy (SBRSS). METHODS: A case series analysis of patients who underwent SBRSS was performed. A surgical robot was used to divide the postganglionic sympathetic fibers and communicating rami to intercostal nerves 2, 3, and 4. The sympathetic chain was left intact. The operation was performed on the dominant side, and the same procedure was then performed on the contralateral side after 4 weeks. The success of the sympathectomy was determined by intraoperative temperature measurement, patient interviews, and the Hyperhidrosis Disease Severity Scale. RESULTS: There were 47 patients (22 men, 25 women), with a mean age of 22 ± 3 years. Minor complications were seen in 4% of patients. One patient had transient heart block. One patient had transient partial Horner's syndrome. Forty percent of patients had transient CH after the first surgery, and 45% had transient CH after sympathectomy on the contralateral side. There were no deaths. The median hospitalization was 3 days. At a mean follow-up of 28 ± 6 months, 46/47 patients (98%) had sustained relief of hyperhidrosis. In one patient (1/47, 2%), hyperhidrosis recurred on the first operated side. One patient (2%) exhibited sustained CH. CONCLUSION: SBRSS is associated with a lower rate of CH than when the procedure is performed bilaterally in a simultaneous fashion. This procedure is associated with the lowest reported rate of CH.

Robotic Lobectomy: Experience with 638 Consecutive Cases.

BACKGROUND: Robotic lobectomy has been evolving over the past decade and has been shown to be an oncologically acceptable procedure. We evaluated our experience with robotic lobectomy for the treatment of early-stage lung cancer. METHODS: We performed a retrospective review of prospectively accrued patients at our institution who underwent robotic lobectomy for early-stage lung cancer from February 2004 to July 2019, RESULTS: Of 3304 consecutive patients who underwent a robotic operation by a single surgeon, 638 underwent robotic lobectomy for early-stage primary non-small cell lung cancer (NSCLC; stages I and II). The 427 (67%) men and 211 (33%) women had a median age of 69 y (range 41-86), and 567 (89 %) were former or current smokers. The median operative time was 176 minutes (range 160-456), the median chest tube time was 3 days (2-8), the median air leak time was 0 days (0-3), and the median length of stay was 3 days (1-26). The median tumor size was 2.6 cm (range 06-3.4). The mean number of nodes recovered was 14 ± 3. Pathologic upstaging was noted in 121 patients (19%). Minor complications were observed in 133 patients (21%). Conversion to thoracotomy occurred in 11 (1.7%) patients. Mortality was 0.5%. CONCLUSION: Robotic lobectomy is a safe, minimally invasive procedure that replicates the oncologic and technical principles of thoracotomy for the treatment of lung cancer.

Robotic First Rib Resection for Thoracic Outlet Syndrome.

PURPOSE: First rib resection is a key component of the treatment of Thoracic Outlet Syndrome (TOS). We report our experience with, and technique for, robotic first rib resection. METHODS: Patients diagnosed with TOS underwent robotic first rib resection of the offending portion of the first rib with disarticulation of the costo-sternal joint. Definitive diagnosis of TOS was made by Magnetic Resonance Angiography (MRA) with maneuvers. RESULTS: A total of 67 patients underwent robotic first rib resection. Neurogenic TOS: 39 patients underwent robotic resection for Neurologic Symptoms of the upper extremity (Neurogenic TOS). There were 14 men and 25 women, with a mean age of 34 ± 9.5 years. Paget-Schroetter Syndrome (PSS) or Venous TOS: 28 patients underwent transthoracic robotic first rib resection for PSS. There were 16 men and 12 women, with a mean age of 24 ± 8.5 years. Operative time was 87.6 ± 10.8 minutes. There were no intraoperative complications. Hospital stay ranged from 2 to 4 days with a median hospitalization of 3 days. There were no neurovascular complications. There was no mortality. In patients with Neurogenic TOS, QuickDASH Scores (mean ± SEM) decreased from 60.3 ± 2.1 preoperatively to 5 ± 2.3 in the immediate postoperative period and to 3.5 ± 1.1 at 6 months (p<0001). Immediate relief of symptoms was seen in 35/39 patients (91%). Persistent paresthesia was seen in 4/39 (9%) immediately postop, and in 2/39 (2.5%) at 6 months. Thirty-seven of 39 (97.5%) patients reported complete relief of symptoms. Among patients with PSS or Venous TOS, 9/28 (32%) required endovascular venoplasty to completely open the subclavian vein after the relief of extrinsic compression. At a median follow-up of 24 months, all patients with PSS had an open subclavian vein, for a patency rate of 100%. CONCLUSIONS: Robotic transthoracic first rib resection allows for minimally invasive resection of the first rib in patients with TOS, with excellent relief of symptoms and no neurovascular complications.

The Effects of Peri-Operative Dexamethasone on Patients Undergoing Total Hip or Knee Arthroplasty: Is It Safe for Diabetics?

BACKGROUND: Peri-operative dexamethasone has been shown to effectively reduce post-operative nausea and vomiting and aide in analgesia after total joint arthroplasty (TJA); however, systemic glucocorticoid therapy has many adverse effects. The purpose of this study is to determine the effects of dexamethasone on prosthetic joint infection (PJI) and blood glucose levels in patients undergoing TJA. METHODS: A retrospective chart review of all patients receiving primary TJA from 2011 to 2015 (n = 2317) was conducted. Patients were divided into 2 cohorts: dexamethasone (n = 1426) and no dexamethasone (n = 891); these groups were subdivided into diabetic and non-diabetic patients. The primary outcome was PJI; secondary measures included glucose levels and pre-operative hemoglobin A1c (A1c) values. Statistics were carried out using logistic and regression models. RESULTS: Of the 2317 joints, 1.12% developed PJI; this was not affected by dexamethasone (P = .166). Diabetics were found to have higher rate of infection (P < .001); however, diabetics who received dexamethasone were not found to have a significantly higher infection rate that non-diabetics (P = .646). Blood glucose levels were found to increase post-operatively, and dexamethasone did not increase this change (P = .537). Diabetes (P < .001) and increasing hemoglobin A1c (P < .001) were also associated with increased serum glucose levels; however, this was not influenced by dexamethasone (P = .595). CONCLUSION: Although diabetic patients were found to have a higher infection rate overall, this was not affected by administration of intravenous dexamethasone, nor was the post-operative elevation in serum glucose levels. In this study population, peri-operative intravenous dexamethasone did not increase the rate of PJI and was safe to administer in patients undergoing TJA.

Long-term Results of Robotic Modified Belsey (Gastroesophageal Valvuloplasty) Fundoplication.

PURPOSE: Nissen fundoplication is associated with poor long-term durability, as well as dysphasia and gas bloat. We report here the long-term results of modified Belsey fundoplication (Gastroesophageal Valvuloplasty; GEV) performed laparoscopically using a surgical robot. METHODS: Patients who underwent robotic GEV were reviewed retrospectively. Operations were performed by laparoscopy and included robotic dissection of the esophageal hiatus, primary closure of the hiatus, followed by intussusception of a 4 cm segment of the esophagus into the stomach for 270°, and suspension of the fundoplication on the hiatal closure. The results were assessed by postoperative endoscopy, contrast esophagography, a Subjective Symptom Questionnaire (SSQ), and objective Visick grading. RESULTS: There were 291 patients (156 male, 135 female, mean age 51±14 years). Indications were intractability (73%) and pulmonary symptoms (27%). Mean operative time was 130 minutes ± 52 minutes. Minor complications were seen in 21%. There was no mortality. Mean hospitalization was 2.8 days ± 1.7 days. Mean follow-up was 85 months ± 7 months. During this period, the mean SSQ score decreased from 8.3 ± 0.6 to 0.7± 0.2 (P < 0.05). There was no long-term dysphasia or gas bloat. Ninety-five percent of patients were Visick I and 5% were Visick II. Hiatal hernia recurred in 7 patients (2%). CONCLUSIONS: Robotic laparoscopic modified Belsey fundoplication (GEV) is associated with excellent long-term durability, reflux control, and low rates of dysphasia and gas bloat. This procedure may represent an alternative to medical antireflux therapy and other surgical antireflux procedures such as Nissen fundoplication.

A kidney-specific genetic control module in mice governs endocrine regulation of the cytochrome P450 gene Cyp27b1 essential for vitamin D3 activation.

The vitamin D endocrine system regulates mineral homeostasis through its activities in the intestine, kidney, and bone. Terminal activation of vitamin D3 to its hormonal form, 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3), occurs in the kidney via the cytochrome P450 enzyme CYP27B1. Despite its importance in vitamin D metabolism, the molecular mechanisms underlying the regulation of the gene for this enzyme, Cyp27b1, are unknown. Here, we identified a kidney-specific control module governed by a renal cell-specific chromatin structure located distal to Cyp27b1 that mediates unique basal and parathyroid hormone (PTH)-, fibroblast growth factor 23 (FGF23)-, and 1,25(OH)2D3-mediated regulation of Cyp27b1 expression. Selective genomic deletion of key components within this module in mice resulted in loss of either PTH induction or FGF23 and 1,25(OH)2D3 suppression of Cyp27b1 gene expression; the former loss caused a debilitating skeletal phenotype, whereas the latter conferred a quasi-normal bone mineral phenotype through compensatory homeostatic mechanisms involving Cyp24a1 We found that Cyp27b1 is also expressed at low levels in non-renal cells, in which transcription was modulated exclusively by inflammatory factors via a process that was unaffected by deletion of the kidney-specific module. These results reveal that differential regulation of Cyp27b1 expression represents a mechanism whereby 1,25(OH)2D3 can fulfill separate functional roles, first in the kidney to control mineral homeostasis and second in extra-renal cells to regulate target genes linked to specific biological responses. Furthermore, we conclude that these mouse models open new avenues for the study of vitamin D metabolism and its involvement in therapeutic strategies for human health and disease.

Extraskeletal Osteosarcoma of the Hand.

Extraskeletal osteosarcoma is a rare disease that uncommonly affects the upper extremity. A 46-year-old African American man presented for evaluation of a right middle finger mass. Excisional biopsy demonstrated extraskeletal osteosarcoma of the soft tissues. We performed a transmetacarpal ray resection.

Epigenetic Plasticity Drives Adipogenic and Osteogenic Differentiation of Marrow-derived Mesenchymal Stem Cells.

Terminal differentiation of multipotent stem cells is achieved through a coordinated cascade of activated transcription factors and epigenetic modifications that drive gene transcription responsible for unique cell fate. Within the mesenchymal lineage, factors such as RUNX2 and PPARγ are indispensable for osteogenesis and adipogenesis, respectively. We therefore investigated genomic binding of transcription factors and accompanying epigenetic modifications that occur during osteogenic and adipogenic differentiation of mouse bone marrow-derived mesenchymal stem cells (MSCs). As assessed by ChIP-sequencing and RNA-sequencing analyses, we found that genes vital for osteogenic identity were linked to RUNX2, C/EBPß, retinoid X receptor, and vitamin D receptor binding sites, whereas adipocyte differentiation favored PPARγ, retinoid X receptor, C/EBPα, and C/EBPß binding sites. Epigenetic marks were clear predictors of active differentiation loci as well as enhancer activities and selective gene expression. These marrow-derived MSCs displayed an epigenetic pattern that suggested a default preference for the osteogenic pathway; however, these patterns were rapidly altered near the Adipoq, Cidec, Fabp4, Lipe, Plin1, Pparg, and Cebpa genes during adipogenic differentiation. Surprisingly, we found that these cells also exhibited an epigenetic plasticity that enabled them to trans-differentiate from adipocytes to osteoblasts (and vice versa) after commitment, as assessed by staining, gene expression, and ChIP-quantitative PCR analysis. The osteogenic default pathway may be subverted during pathological conditions, leading to skeletal fragility and increased marrow adiposity during aging, estrogen deficiency, and skeletal unloading. Taken together, our data provide an increased mechanistic understanding of the epigenetic programs necessary for multipotent differentiation of MSCs that may prove beneficial in the development of therapeutic strategies.

Cost of physician-led home visit care (Zaitaku care) compared with hospital care at the end of life in Japan.

BACKGROUND: Physician-led home visit care with medical teams (Zaitaku care) has been developed on a national scale to support those who wish to stay at home at the end of life, and promote a system of community-based integrated care in Japan. Medical care at the end of life can be expensive, and is an urgent socioeconomic issue for aging societies. However medical costs of physician-led home visits care have not been well studied. We compared the medical costs of Zaitaku care and hospital care at the end of life in a rapidly aging community in a rural area in Japan. METHODS: A cross-sectional study was performed to compare the total medical costs during patients' final days of life (30 days or less) between Zaitaku care and hospital care from September 2012 to August 2013 in Fukuoka Prefecture, Japan. RESULTS: Thirty four patients died at home under Zaitaku care, and 72 patients died in the hospital during this period. The average daily cost of care during the last 30 days did not differ significantly between the two groups. Although Zaitaku care costs were higher than hospital care costs in the short-term (≦10 days, Zaitaku care $371.2 vs. Hospital care $202.0, p = 0.492), medical costs for Zaitaku care in the long-term care (≧30 days) were less than that of hospital care ($155.8 vs. $187.4, p = 0.055). CONCLUSIONS: Medical costs of Zaitaku care were less compared with hospital care if incorporated early for long term care, but it was high if incorporated late for short term care. For long term care, medical costs for Zaitaku care was 16.7% less than for hospitalization at the end of life. This physician-led home visit care model should be an available option for patients who wish to die at home, and may be beneficial financially over time.

Smartphone-Based Goniometry Accuracy in Clinical Scenarios.

This study compared two popular iPhone-based goniometer applications to the gold standard universal goniometer for the measurement of the hip and knee joints in scenarios mimicking the normal pace of an orthopaedic clinical practice.Three physicians measured hip and knee joint angles 35 times with one of three goniometers: universal 12-inch goniometer, DrGoniometer (iPhone-5 based), and SimpleGoniometer (iPhone-5 based). Data wwere analyzed using Pearson correlation coefficient calculations. Average knee angles measured with the universal goniometer, DrGoniometer, and SimpleGoniometer measured 83.46°, 85.23°, and 80.39°, respectively. The smartphone-based goniometers had moderate agreement with the universal goniometer in the knee (r > .322). Average hip angles measured with the universal goniometer, DrGoniometer, and SimpleGoniometer measured 62.34°, 60.87°, and 59.34°, respectively. The smartphone-based goniometers had moderate agreement with the universal goniometer in the hip (r > .168). Smartphone-based goniometers gave accurate, with weak to moderate correlation, measurements for the knee and hip. (Journal of Surgical Orthopaedic Advances 26(4):223-226, 2017).

Selective Distal Enhancer Control of the Mmp13 Gene Identified through Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) Genomic Deletions.

Matrix metalloproteinase 13 (Mmp13, collagenase-3) plays an essential role in bone metabolism and mineral homeostasis. It is regulated by numerous factors, including BMP-2, parathyroid hormone, and 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3), through transcription factors such as Runt-related transcription factor 2 (RUNX2), CCAAT/enhancer-binding protein ß (C/EBPß), OSX, and vitamin D receptor (VDR). During osteoblast maturation, the basal expression of Mmp13 and its sensitivity to 1,25(OH)2D3 are strikingly increased. In this report, ChIP-sequencing analysis in mouse preosteoblasts revealed that the Mmp13 gene was probably regulated by three major enhancers located -10, -20, and -30 kb upstream of the gene promoter, occupied by activated VDR and prebound C/EBPß and RUNX2, respectively. Initially, bacterial artificial chromosome clone recombineering and traditional mutagenesis defined binding sites for VDR and RUNX2. We then employed a CRISPR/Cas9 gene editing approach to delete the -10 and -30 kb Mmp13 enhancers, a region proximal to the promoter, and VDR or RUNX2. VDR-mediated up-regulation of Mmp13 transcription was completely abrogated upon removal of the -10 kb enhancer, resulting in a 1,25(OH)2D3-directed repression of Mmp13. Deletion of either the -30 kb enhancer or RUNX2 resulted in a complete loss of basal transcript activity and a ChIP-identified destabilization of the chromatin enhancer environment and factor binding. Whereas enhancer deletions only affected Mmp13 expression, the RUNX2 deletion led to changes in gene expression, a reduction in cellular proliferation, and an inability to differentiate. We conclude that the Mmp13 gene is regulated via at least three specific distal enhancers that display independent activities yet are able to integrate response from multiple signaling pathways in a model of activation and suppression.

Mechanisms of Enhancer-mediated Hormonal Control of Vitamin D Receptor Gene Expression in Target Cells.

The biological actions of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) are mediated by the vitamin D receptor (VDR), whose expression in bone cells is regulated positively by 1,25(OH)2D3, retinoic acid, and parathyroid hormone through both intergenic and intronic enhancers. In this report, we used ChIP-sequencing analysis to confirm the presence of these Vdr gene enhancers in mesenchyme-derived bone cells and to describe the epigenetic histone landscape that spans the Vdr locus. Using bacterial artificial chromosome-minigene stable cell lines, CRISPR/Cas9 enhancer-deleted daughter cell lines, transient transfection/mutagenesis analyses, and transgenic mice, we confirmed the functionality of these bone cell enhancers in vivo as well as in vitro. We also identified VDR-binding sites across the Vdr gene locus in kidney and intestine using ChIP-sequencing analysis, revealing that only one of the bone cell-type enhancers bound VDR in kidney tissue, and none were occupied by the VDR in the intestine, consistent with weak or absent regulation by the 1,25(OH)2D3 hormone in these tissues, respectively. However, a number of additional sites of VDR binding unique to either kidney or intestine were present further upstream of the Vdr gene, suggesting the potential for alternative regulatory loci. Importantly, virtually all of these regions retained histone signatures consistent with those of enhancers and exhibited unique DNase I hypersensitivity profiles that reflected the potential for chromatin access. These studies define mechanisms associated with hormonal regulation of the Vdr and hint at the differential nature of VDR binding activity at the Vdr gene in different primary target tissues in vivo.