Association between vitamin D status and eryptosis-results from the German National Cohort Study.

Vitamin D, besides its classical effect on mineral homeostasis and bone remodeling, can also modulate apoptosis. A special form of apoptosis termed eryptosis appears in erythrocytes. Eryptosis is characterized by cell shrinkage, membrane blebbing, and cell membrane phospholipid disorganization and associated with diseases such as sepsis, malaria or iron deficiency, and impaired microcirculation. To our knowledge, this is the first study that linked vitamin D with eryptosis in humans. This exploratory cross-sectional trial investigated the association between the vitamin D status assessed by the concentration of plasma 25-hydroxyvitamin D (25(OH)D) and eryptosis. Plasma 25(OH)D was analyzed by LC-MS/MS, and eryptosis was estimated from annexin V-FITC-binding erythrocytes by FACS analysis in 2074 blood samples from participants of the German National Cohort Study. We observed a weak but clear correlation between low vitamin D status and increased eryptosis (r = - 0.15; 95% CI [- 0.19, - 0.10]). There were no differences in plasma concentrations of 25(OH)D and eryptosis between male and female subjects. This finding raises questions of the importance of vitamin D status for eryptosis in terms of increased risk for anemia or cardiovascular events.

Microalgae-derived sterols do not reduce the bioavailability of oral vitamin D3 in mice.

Microalgae have drawn increasing attention as sustainable food sources, also because of their lipid-lowering phytosterols. As phytosterols are also discussed critically regarding their effect on the availability of fat-soluble vitamins, this study aimed to investigate microalgae-derived phytosterols and their effect on vitamin D status. GC-MS analysis showed large variations in the phytosterol profiles of microalgal species. The most frequent sterols were ß-sitosterol and stigmasterol. To investigate their effects on vitamin D status, 40 mice were randomized to four groups and fed a vitamin D3-adequate (25 µg/kg) Western-style diet with 0% phytosterols (control) or 1% ergosterol (a fungal sterol not typical for microalgae), ß-sitosterol or stigmasterol for four weeks. Contrary to the hypothesis that phytosterols adversely affect vitamin D uptake, mice fed ß-sitosterol had significantly higher concentrations of vitamin D3 in plasma (3.15-fold, p<0.01), liver (3.15-fold, p<0.05), and skin (4.12-fold, p<0.005) than the control group. Small increases in vitamin D3 in plasma and skin were also observed in mice fed stigmasterol. In contrast, vitamin D3 levels in the ergosterol and control groups did not differ. The increased tissue levels of vitamin D3 in mice fed ß-sitosterol and stigmasterol were not attributable to the observed reduction in liver triglycerides in these groups. The data rather suggest that changes in bile acid profiles were responsible for the beneficial effect of microalgae sterols on the bioavailability of vitamin D3. In conclusion, consumption of microalgae might not adversely affect vitamin D status.

Genotype-dependent impact of dietary vitamin D3 on laying hens.

Vitamin D3 has an integral part in calcium and phosphorus homoeostasis, which in turn plays a key role in egg production of hens. The present study aimed to investigate whether an additional vitamin D3 supplementation improves the laying performance and egg quality of hens according to their genetic potential. For this purpose, four layer lines (low performing: R11 and L68; high performing: WLA and BLA) supplemented either with 300 or 3000 IU vitamin D3 per kg feed were compared concerning serum 25-hydroxyvitamin D3 (25-OHD3), calcium, phosphorus and alkaline phosphatase (ALP), laying performance and egg quality. The higher supplementation of vitamin D3 increased 25-OHD3 serum concentrations in all genotypes, except for R11 and WLA hens in week 49, and also elevated vitamin D3 and 25-OHD3 content in the egg yolk (p < 0.05). In week 29, 3000 IU vitamin D3 decreased pooled least squares means (LSMeans) of serum calcium concentrations considering all genotypes and increased the ALP concentrations in BLA hens (p < 0.05). Considering the whole experimental period daily egg mass of R11 hens was increased by an additional vitamin D3 supplementation (p < 0.001). Regarding all genotypes and the whole experimental period the pooled LSMeans of breaking strength of eggs from hens fed 3000 IU vitamin D3 were higher than those of hens fed 300 IU (p = 0.044). In conclusion, present results give evidence that the higher vitamin D3 supplementation might have genotype-dependently beneficial effects on calcium and phosphorus homoeostasis of hens, which might improve feed efficiency in the early laying period and promote the persistence of the laying period irrespectively of genotype. The increase of serum 25-OHD3 by the higher vitamin D supplementation supported the higher transfer of vitamin D in the egg yolk and improved genotype-dependently the breaking strength of the eggshell.

Prevalence of vitamin D insufficiency and evidence for disease prevention in the older population.

The prevalence of vitamin D insufficiency, usually assessed by the analysis of circulating 25-hydroxyvitamin D (25[OH])D), is very high in the aging German population. An important factor that reduces endogenous vitamin D synthesis in older persons is physical inactivity or care-dependency that limits the time spent outside. Additionally, it has been suggested that the age-dependent decline in the glomerular filtration rate is associated with a reduced production of bioactive calcitriol. As this phenomenon is not detectable by the assessment of 25(OH)D, it is necessary to analyze the level of parathyroid hormone as a marker of calcitriol function. Because 25(OH)D levels are highly correlated with an active and healthy life style, data from epidemiological studies are not necessarily suitable to elucidate the role of vitamin D in disease prevention. Recently published meta-analyses of randomized controlled trials (RCTs) showed moderate effects of vitamin D supplementation on fracture risk and found that vitamin D was more effective when administered in combination with calcium. The role of vitamin D in the prevention of falls and frailty remains unclear. Much evidence has demonstrated the beneficial effects of vitamin D on respiratory tract infections and asthma, which are very relevant health issues in the older population. To conclude, vitamin D, particularly combined with calcium, has moderately beneficial effects on the skeletal system and is useful for the prevention of respiratory tract infections.

Functional Synaptic Integration of Forebrain GABAergic Precursors into the Adult Spinal Cord.

Spinal cord transplants of embryonic cortical GABAergic progenitor cells derived from the medial ganglionic eminence (MGE) can reverse mechanical hypersensitivity in the mouse models of peripheral nerve injury- and paclitaxel-induced neuropathic pain. Here, we used electrophysiology, immunohistochemistry, and electron microscopy to examine the extent to which MGE cells integrate into host circuitry and recapitulate endogenous inhibitory circuits. Whether the transplants were performed before or after nerve injury, the MGE cells developed into mature neurons and exhibited firing patterns characteristic of subpopulations of cortical and spinal cord inhibitory interneurons. Conversely, the transplanted cells preserved cortical morphological and neurochemical properties. We also observed a robust anatomical and functional synaptic integration of the transplanted cells into host circuitry in both injured and uninjured animals. The MGE cells were activated by primary afferents, including TRPV1-expressing nociceptors, and formed GABAergic, bicuculline-sensitive, synapses onto host neurons. Unexpectedly, MGE cells transplanted before injury prevented the development of mechanical hypersensitivity. Together, our findings provide direct confirmation of an extensive, functional synaptic integration of MGE cells into host spinal cord circuits. This integration underlies normalization of the dorsal horn inhibitory tone after injury and may be responsible for the prophylactic effect of preinjury transplants. SIGNIFICANCE STATEMENT: Spinal cord transplants of embryonic cortical GABAergic interneuron progenitors from the medial ganglionic eminence (MGE), can overcome the mechanical hypersensitivity produced in different neuropathic pain models in adult mice. Here, we examined the properties of transplanted MGE cells and the extent to which they integrate into spinal cord circuitry. Using electrophysiology, immunohistochemistry, and electron microscopy, we demonstrate that MGE cells, whether transplanted before or after nerve injury, develop into inhibitory neurons, are activated by nociceptive primary afferents, and form GABA-A-mediated inhibitory synapses with the host. Unexpectedly, cells transplanted into naive spinal cord prevented the development of nerve-injury-induced mechanical hypersensitivity. These results illustrate the remarkable plasticity of adult spinal cord and the potential of cell-based therapies against neuropathic pain.

Impact of chocolate liquor on vascular lesions in apoE-knockout mice.

Cocoa polyphenols are thought to reduce the risk of cardiovascular diseases. Thus, cocoa-containing foods may have significant health benefits. Here, we studied the impact of chocolate liquor on vascular lesion development and plaque composition in a mouse model of atherosclerosis. Apolipoprotein E (apoE)-knockout mice were assigned to two groups and fed a Western diet that contained 250 g/kg of either chocolate liquor or a polyphenol-free isoenergetic control paste for 16 weeks. In addition to fat, protein, and fibers, the chocolate liquor contained 2 g/kg of polyphenols. Compared with the control group, mice fed the chocolate liquor had larger plaque areas in the descending aorta and aortic root, which were attributed to a higher mass of vascular smooth muscle cells (VSMCs) and collagen. Vascular lipid deposits and calcification areas did not differ between the two groups. The aortic tissue level of interleukin-6 (IL-6) mRNA was 5-fold higher in the mice fed chocolate liquor than in the control mice. Chocolate-fed mice exhibited an increased hepatic saturated to polyunsaturated fatty acid ratio than the controls. Although the chocolate liquor contained 14 µg/kg of vitamin D2, the chocolate liquor-fed mice did not have measurable 25-hydroxyvitamin D2 in the serum. These mice even showed a 25% reduction in the level of 25-hydroxyvitamin D3 compared with the control mice. Overall, present data may contribute to our understanding how chocolate constituents can impact vascular lesion development.

The prognostic effect of high diagnostic WT1 gene expression in pediatric AML depends on WT1 SNP rs16754 status: report from the Children's Oncology Group.

BACKGROUND: WT1 is aberrantly over-expressed in most cases of AML. We recently demonstrated that WT1 SNP rs16754 correlates with favorable outcome and high diagnostic WT1 expression in childhood AML. We examined the clinical correlates of diagnostic WT1 expression within a contemporary COG trial and determined whether its prognostic impact differs between SNP+ and SNP- patients. PROCEDURE: WT1 mRNA expression was measured via qRT-PCR in diagnostic specimens obtained from 225 patients enrolled on COG-AAML03P1. Direct sequencing of WT1 exon 7 was performed to determine SNP rs16754 genotype. WT1 expression was correlated with disease characteristics, SNP status, and outcome. RESULTS: Patients were categorized into four groups (quartiles: Q1 through Q4) based on diagnostic WT1 expression for analysis. FLT3/ITD (P = 0.017) and WT1 mutations (P < 0.001) both occurred more frequently in patients with the highest WT1 expression. SNP rs16754 frequency did not vary significantly among the quartiles. When all patients were considered, survival outcomes were similar between quartiles. However, when only SNP- patients (n = 150) were analyzed, those with highest WT1 expression (Q4) had the poorest OS (51% vs. 72% for Q1-Q3, P = 0.006) and EFS (35% vs. 54% for Q1-Q3, P = 0.031). Among SNP+ patients (n = 75), survival did not vary significantly between WT1 expression quartiles. CONCLUSION: Although WT1 expression was not prognostic when all patients were considered together, stratifying patients by SNP rs16754 genotype revealed significant differences in outcome. In SNP- patients, high WT1 expression predicted decreased survival in univariate, but not multivariate, analysis, due to a preponderance of high-risk cyto/molecular abnormalities in the highest expression quartile.

Addressing psychosocial needs in patients with Long-COVID (PsyLoCo-Study): study protocol of a pilot-study of a specialized modular intervention.

Introduction: After an acute infection with the corona virus 10-20% of those affected suffer from ongoing or new symptoms. A causal therapy for the phenomenon known as Long/Post-COVID is still lacking and specific therapies addressing psychosocial needs of these patients are imperatively needed. The aim of the PsyLoCo-study is developing and piloting a psychotherapeutic manual, which addresses Long/Post-COVID-related psychosocial needs and supports in coping with persistent bodily symptoms as well as depressive or anxiety symptoms. Methods and analysis: This pilot trial implements a multi-centre, 2-arm (N=120; allocation ratio: 1:1), parallel group, randomised controlled design. The pilot trial is designed to test the feasibility and estimate the effect of 1) a 12-session psychotherapeutic intervention compared to 2) a wait-list control condition on psychosocial needs as well as bodily and affective symptoms in patients suffering from Long/Post-COVID. The intervention uses an integrative, manualized, psychotherapeutic approach. The primary study outcome is health-related quality of life. Outcome variables will be assessed at three timepoints, pre-intervention (t1), post-intervention (t2) and three months after completed intervention (t3). To determine the primary outcome, changes from t1 to t2 are examined. The analysis will be used for the planning of the RCT to test the efficacy of the developed intervention. Discussion: The pilot study will evaluate a 12-session treatment manual for Long/Post-COVID sufferers and the therapy components it contains. The analysis will provide insights into the extent to which psychotherapeutic treatment approaches improve the symptoms of Long/Post-COVID sufferers. The treatment manual is designed to be carried out by psychotherapists as well as people with basic training in psychotherapeutic techniques. This approach was chosen to enable a larger number of practitioners to provide therapeutic support for Long/Post-COVID patients. After completion of the pilot study, it is planned to follow up with a randomized controlled study and to develop a treatment guideline for general practitioners and interested specialists. Trial registration: The pilot trial has been registered with the German Clinical Trials Register (Deutsches Register Klinischer Studien; Trial-ID: DRKS00030866; URL: https://drks.de/search/de/trial/DRKS00030866) on March 7, 2023.

Liquid biopsy for improving diagnosis and monitoring of CNS lymphomas: A RANO review.

BACKGROUND: The utility of liquid biopsies is well documented in several extracranial and intracranial (brain/leptomeningeal metastases, gliomas) tumors. METHODS: The RANO (Response Assessment in Neuro-Oncology) group has set up a multidisciplinary Task Force to critically review the role of blood and cerebrospinal fluid (CSF)-liquid biopsy in CNS lymphomas, with a main focus on primary central nervous system lymphomas (PCNSL). RESULTS: Several clinical applications are suggested: diagnosis of PCNSL in critical settings (elderly or frail patients, deep locations, and steroid responsiveness), definition of minimal residual disease, early indication of tumor response or relapse following treatments, and prediction of outcome. CONCLUSIONS: Thus far, no clinically validated circulating biomarkers for managing both primary and secondary CNS lymphomas exist. There is need of standardization of biofluid collection, choice of analytes, and type of technique to perform the molecular analysis. The various assays should be evaluated through well-organized central testing within clinical trials.

Prognostic implications of the IDH1 synonymous SNP rs11554137 in pediatric and adult AML: a report from the Children's Oncology Group and SWOG.

IDH1 SNP rs11554137 was recently reported in association with poor prognosis in normal karyotype adult acute myeloid leukemia (AML). We aimed to determine the prevalence, clinical associations, and prognostic significance of SNP rs11554137 in unselected pediatric and adult AML patients. Diagnostic marrow specimens from 527 AML patients treated on the pediatric trial Children's Oncology Group-AAML03P1 (N = 253) or adult SWOG trials (N = 274) were analyzed for the presence of the SNP. SNP rs11554137 was present in 11% of all patients. SNP status had no prognostic impact on survival in pediatric patients. In adult AML, overall survival for SNP-positive patients was 10% versus 18% for SNP-negative patients (P = .44). Among the 142 adults who achieved complete remission, 5-year relapse-free survival was significantly worse for SNP-positive patients (0% vs 25%, P = .0014). However, among adults with normal cytogenetics, FLT3/ITD was present in 90% of SNP-positive patients versus 59% of SNP-negative patients (P = .0053). In multivariate analysis, adjusting for the effects of age, cytogenetics, and FLT3/ITD, the independent prognostic effect of SNP positivity was not statistically significant (hazard ratio = 1.72, P = .18). The clinical profile of SNP-positive patients suggests that SNP rs11554137 may have biologic effects that bear further investigation. The clinical trials in this study are registered at http://www.clinicaltrials.gov as #NCT000707174 and #NCT00899171.

Knowledge, Attitudes and Behaviors regarding Tick-Borne Encephalitis Vaccination and Prevention of Tick-Borne Diseases among Primary Care Physicians in Bavaria and Baden-Wuerttemberg, Germany, May-September 2022.

In 2020, a record number of tick-borne encephalitis (TBE) cases was reported in major endemic areas in Germany, i.e., the southern federal states of Baden-Wuerttemberg and Bavaria. Most cases were unvaccinated. Other tick-borne diseases (TBDs), including Lyme borreliosis and tularemia, are rising, too. Thus, strategies are needed to increase TBE vaccination uptake in risk areas and promote education on TBD prevention. Primary care physicians are key providers of both vaccinations and TBD education. The TBD-Prevention (TBD-Prev) study aimed to investigate the knowledge, attitudes and behaviors of primary care physicians in Baden-Wuerttemberg and Bavaria with regard to TBE vaccination and prevention of TBDs and to derive strategies for increasing vaccination rates and improving knowledge about TBE and other TBDs in the population and among primary care physicians. We invited all primary care physicians (N = 14,046) in both states to participate by mail. Using standardized, self-administered questionnaires, available both on paper and online, we asked physicians anonymously about their knowledge, attitudes and behaviors with respect to TBE vaccination and TBD prevention and their need for further information/educational materials. A total of 2321 physicians participated between May and September 2022 (response rate 17%), of whom 1222 (53%) worked in Baden-Wuerttemberg and 1067 (46%) in Bavaria. Among the participating physicians, 56% were male, 71% were >50 years and 51% worked in an individual practice. Furthermore, 91% were aware of the German national vaccination guidelines, and 98% perceived their knowledge of the risks and benefits of vaccination as adequate. A total of 97% offer TBE vaccinations, 67% provide vaccination counselling during initial consultations with new patients and 64% actively remind patients about due vaccinations. In addition, 24% expressed a need for further information materials, mainly traditional, analogue media such as flyers (82%) and posters (50%), and named timeliness, quality assurance, easy comprehensibility and independence from the pharmaceutical industry as the most important characteristics of such materials. Almost all participating physicians reported offering TBE vaccinations and feeling well-informed about TBE vaccination and TBDs. However, active offering of vaccinations and education could be further improved, and additional, low-threshold information materials are needed. Based on these results, we will develop and provide various materials on TBE vaccination and TBDs, in particular flyers and posters, for use by physicians during consultations.

Impact of Regular Intake of Microalgae on Nutrient Supply and Cardiovascular Risk Factors: Results from the NovAL Intervention Study.

A 14-day randomized controlled study with a parallel design was conducted with 80 healthy participants. Intervention groups I (IG1) and II (IG2) received a defined background diet and consumed a smoothie enriched with either 15 g of Chlorella dry weight (d.w.) or 15 g of Microchloropsis d.w. daily. Control group II (CG2) received a defined background diet without the smoothie. Control group I (CG1) received neither. Blood samples and 24-h urine were collected at the beginning and the end of the study. Serum concentrations of 25-hydroxyvitamin D3, vitamin D3, selenium, iron, ferritin, transferrin saturation, total cholesterol, low-density lipoprotein (LDL) cholesterol, high-density lipoprotein (HDL) cholesterol, non-HDL cholesterol and the LDL-cholesterol/HDL cholesterol ratio decreased in IG1 (p < 0.05), while 25-hydroxyvitamin D2 increased (p < 0.05). In IG2, vitamin D3, 25-hydroxyvitamins D2 and D3 decreased (p < 0.05), while concentrations of fatty acids C20:5n3 and C22:5n3 increased. Serum and urine uric acid increased in IG1 and IG2 (p < 0.05). Microchloropsis is a valuable source of n3 fatty acids, as is Chlorella of vitamin D2. Regular consumption of Chlorella may affect the iron and selenium status negatively but may impact blood lipids positively. An elevated uric acid concentration in blood and urine following the regular consumption of microalgae poses potential risks for human health.

Time since death: The histological chronotanatognostic.

The time since death is an important aspect of forensic medicine; however, there is not an accurate single method to determine this data. Therefore, this research aimed to evaluate parameters and procedures based on the morphological analysis of cells and tissues to determine the time since death, using animal models. Pigs were chosen in this research because of their similarities with human anatomy, physiology, and pathophysiology. We identified the cells and tissue alterations in the viscera of pig cadavers according to the time since death, also describing the changes in the temperature of the organs and the bodies. The environmental temperature during the sample collection was also registered. The viscera analysis was performed for 24 h, with a 2-h variation period. After the sample collection, microscope slides were prepared for optical microscopy analysis. Through this 24-h analysis, we observed that the pancreas, small intestine, and large intestine presented more cellular alterations than the other organs. The alterations observed in the other viscera have significance when analyzed in combination. The meninges presented higher stability and few changes in 24 h, which could be relevant in an investigation of the time since death in a period greater than 24 h. Our results showed that histological evaluation is an excellent method to determine the time since death.

Comparison of Mid-Infrared Handheld and Benchtop Spectrometers to Detect Staphylococcus epidermidis in Bone Grafts.

Bone analyses using mid-infrared spectroscopy are gaining popularity, especially with handheld spectrometers that enable on-site testing as long as the data quality meets standards. In order to diagnose Staphylococcus epidermidis in human bone grafts, this study was carried out to compare the effectiveness of the Agilent 4300 Handheld Fourier-transform infrared with the Perkin Elmer Spectrum 100 attenuated-total-reflectance infrared spectroscopy benchtop instrument. The study analyzed 40 non-infected and 10 infected human bone samples with Staphylococcus epidermidis, collecting reflectance data between 650 cm-1 and 4000 cm-1, with a spectral resolution of 2 cm-1 (Agilent 4300 Handheld) and 0.5 cm-1 (Perkin Elmer Spectrum 100). The acquired spectral information was used for spectral and unsupervised classification, such as a principal component analysis. Both methods yielded significant results when using the recommended settings and data analysis strategies, detecting a loss in bone quality due to the infection. MIR spectroscopy provides a valuable diagnostic tool when there is a tissue shortage and time is of the essence. However, it is essential to conduct further research with larger sample sizes to verify its pros and cons thoroughly.

Circulating Tumor DNA Profiling for Detection, Risk Stratification, and Classification of Brain Lymphomas.

PURPOSE: Clinical outcomes of patients with CNS lymphomas (CNSLs) are remarkably heterogeneous, yet identification of patients at high risk for treatment failure is challenging. Furthermore, CNSL diagnosis often remains unconfirmed because of contraindications for invasive stereotactic biopsies. Therefore, improved biomarkers are needed to better stratify patients into risk groups, predict treatment response, and noninvasively identify CNSL. PATIENTS AND METHODS: We explored the value of circulating tumor DNA (ctDNA) for early outcome prediction, measurable residual disease monitoring, and surgery-free CNSL identification by applying ultrasensitive targeted next-generation sequencing to a total of 306 tumor, plasma, and CSF specimens from 136 patients with brain cancers, including 92 patients with CNSL. RESULTS: Before therapy, ctDNA was detectable in 78% of plasma and 100% of CSF samples. Patients with positive ctDNA in pretreatment plasma had significantly shorter progression-free survival (PFS, P < .0001, log-rank test) and overall survival (OS, P = .0001, log-rank test). In multivariate analyses including established clinical and radiographic risk factors, pretreatment plasma ctDNA concentrations were independently prognostic of clinical outcomes (PFS HR, 1.4; 95% CI, 1.0 to 1.9; P = .03; OS HR, 1.6; 95% CI, 1.1 to 2.2; P = .006). Moreover, measurable residual disease detection by plasma ctDNA monitoring during treatment identified patients with particularly poor prognosis following curative-intent immunochemotherapy (PFS, P = .0002; OS, P = .004, log-rank test). Finally, we developed a proof-of-principle machine learning approach for biopsy-free CNSL identification from ctDNA, showing sensitivities of 59% (CSF) and 25% (plasma) with high positive predictive value. CONCLUSION: We demonstrate robust and ultrasensitive detection of ctDNA at various disease milestones in CNSL. Our findings highlight the role of ctDNA as a noninvasive biomarker and its potential value for personalized risk stratification and treatment guidance in patients with CNSL.[Media: see text].

Ca++/CaMKII switches nociceptor-sensitizing stimuli into desensitizing stimuli.

Many extracellular factors sensitize nociceptors. Often they act simultaneously and/or sequentially on nociceptive neurons. We investigated if stimulation of the protein kinase C epsilon (PKCε) signaling pathway influences the signaling of a subsequent sensitizing stimulus. Central in activation of PKCs is their transient translocation to cellular membranes. We found in cultured nociceptive neurons that only a first stimulation of the PKCε signaling pathway resulted in PKCε translocation. We identified a novel inhibitory cascade to branch off upstream of PKCε, but downstream of Epac via IP3-induced calcium release. This signaling branch actively inhibited subsequent translocation and even attenuated ongoing translocation. A second 'sensitizing' stimulus was rerouted from the sensitizing to the inhibitory branch of the signaling cascade. Central for the rerouting was cytoplasmic calcium increase and CaMKII activation. Accordingly, in behavioral experiments, activation of calcium stores switched sensitizing substances into desensitizing substances in a CaMKII-dependent manner. This mechanism was also observed by in vivo C-fiber electrophysiology corroborating the peripheral location of the switch. Thus, we conclude that the net effect of signaling in nociceptors is defined by the context of the individual cell's signaling history.

Integrated Intraoperative Optical Coherence Tomography during Scleral Pass in Strabismus Surgery.

We describe a novel application of integrated intraoperative OCT (iiOCT) to strabismus surgery during the scleral pass and demonstrate it to be a useful tool. A number of complications can arise from inappropriate scleral pass depth during strabismus surgery, leading to an increased risk of unwanted complications including endophthalmitis, retinal detachment, and a lost or slipped muscle. Our study demonstrated that the use of iiOCT provides easy to interpret, real-time feedback to the strabismus surgeon and may translate to safer, more consistent scleral suturing during strabismus surgery and strabismus surgical training. .

Estrogen destabilizes microtubules through an ion-conductivity-independent TRPV1 pathway.

Recently, we described estrogen and agonists of the G-protein coupled estrogen receptor GPR30 to induce protein kinase C (PKC)ε-dependent pain sensitization. PKCε phosphorylates the ion channel transient receptor potential, vanilloid subclass I (TRPV1) close to a novel microtubule-TRPV1 binding site. We now modeled the binding of tubulin to the TRPV1 C-terminus. The model suggests PKCε phosphorylation of TRPV1-S800 to abolish the tubulin-TRPV1 interaction. Indeed, in vitro PKCε phosphorylation of TRPV1 hindered tubulin-binding to TRPV1. In vivo, treatment of sensory neurons and F-11 cells with estrogen and the GPR30 agonist, G-1, resulted in microtubule destabilization and retraction of microtubules from filopodial structures. We found estrogen and G-1 to regulate the stability of the microtubular network via PKC phosphorylation of the PKCε-phosphorylation site TRPV1-S800. Microtubule disassembly was not, however, dependent on TRPV1 ion conductivity. TRPV1 knock-down in rats inverted the effect of the microtubule-modulating drugs, Taxol and Nocodazole, on estrogen-induced and PKCε-dependent mechanical pain sensitization. Thus, we suggest the C-terminus of TRPV1 to be a signaling intermediate downstream of estrogen and PKCε, regulating microtubule-stability and microtubule-dependent pain sensitization.

Oral Intake of Lumisterol Affects the Metabolism of Vitamin D.

SCOPE: The treatment of food with ultraviolet-B (UV-B) light to increase the vitamin D content is accompanied by the formation of photoisomers, such as lumisterol2 . The physiological impact of photoisomers is largely unknown. METHODS AND RESULTS: Three groups of C57Bl/6 mice are fed diets containing 50 µg kg-1 deuterated vitamin D3 with 0, 50 (moderate-dose) or 2000 µg kg-1 (high-dose) lumisterol2 for four weeks. Considerable quantities of lumisterol2 and vitamin D2 are found in the plasma and tissues of mice fed with 2000 µg kg-1 lumisterol2 but not in those fed 0 or 50 µg kg-1 lumisterol2 . Mice fed with 2000 µg kg-1 lumisterol2 showed strongly reduced deuterated 25-hydroxyvitamin D3 (-50%) and calcitriol (-80%) levels in plasma, accompanied by downregulated mRNA abundance of cytochrom P450 (Cyp)27b1 and upregulated Cyp24a1 in the kidneys. Increased tissue levels of vitamin D2 were also seen in mice in a second study that are kept on a diet with 0.2% UV-B exposed yeast versus those fed 0.2% untreated yeast containing iso-amounts of vitamin D2 . CONCLUSION: High doses of lumisterol2 can enter the body, induce the formation of vitamin D2 , reduce the levels of 25(OH)D3 and calcitriol and strongly impact the expression of genes involved in the degradation and synthesis of bioactive vitamin D.

Regulatory T-cells inhibit microglia-induced pain hypersensitivity in female mice.

Peripheral nerve injury-induced neuropathic pain is a chronic and debilitating condition characterized by mechanical hypersensitivity. We previously identified microglial activation via release of colony-stimulating factor 1 (CSF1) from injured sensory neurons as a mechanism contributing to nerve injury-induced pain. Here, we show that intrathecal administration of CSF1, even in the absence of injury, is sufficient to induce pain behavior, but only in male mice. Transcriptional profiling and morphologic analyses after intrathecal CSF1 showed robust immune activation in male but not female microglia. CSF1 also induced marked expansion of lymphocytes within the spinal cord meninges, with preferential expansion of regulatory T-cells (Tregs) in female mice. Consistent with the hypothesis that Tregs actively suppress microglial activation in females, Treg deficient (Foxp3DTR) female mice showed increased CSF1-induced microglial activation and pain hypersensitivity equivalent to males. We conclude that sexual dimorphism in the contribution of microglia to pain results from Treg-mediated suppression of microglial activation and pain hypersensitivity in female mice.