Tight correlation of 5-hydroxymethylcytosine and Polycomb marks in health and disease.

Modifications to DNA and histone tails represent key epigenetic marks involved in establishing and maintaining cell identity and can be dysregulated in human diseases, including cancer. Two such modifications, tri-methylation of lysine-27 on histone H3 (H3K27me3) mediated by the Polycomb complex and hydroxymethylation of cytosines on DNA, have recently been shown to be dynamically regulated during differentiation. Here, we show that global levels of 5-hydroxymethylcytosine (5hmC) and H3K27me3 are highly correlated across a variety of somatic tissues. In multiple hierarchically organized tissues, both marks showed almost identical cell-by-cell distribution patterns that exhibited a tight association with differentiation. In particular, tissue stem cell compartments were characterized by low levels of both marks, whereas differentiated cell compartments exhibited high levels of 5hmC and H3K27me3. This pattern of correlation between the two marks could be recapitulated in an in vitro model system of induced differentiation in prostate epithelial cells. While the correlation between 5hmC and H3K27me3 levels is also maintained in human cancers, the degree of correlation is reduced. These findings suggest a previously unappreciated link between 5hmC and H3K27me3 regulation that should be explored in future mechanistic studies.

Is there a correlation between structural alterations and retinal sensitivity in morphological patterns of diabetic macular edema?

Spectral domain optical coherence tomography (SDOCT) enables enhanced visualization of retinal layers and delineation of structural alterations in diabetic macular edema (DME). Microperimetry (MP) is a new technique that allows fundus-related testing of local retinal sensitivity. Combination of these two techniques would enable a structure-function correlation with insights into pathomechanism of vision loss in DME. To correlate retinal structural derangement with retinal sensitivity alterations in cases with diabetic macular edema, using SDOCT and MP. Prospective study of 34 eyes of 30 patients with DME. All patients underwent comprehensive ophthalmic examination, fluorescein angiography, microperimetry and SDOCT. Four distinct morphological patterns of DME were identified- diffuse retinal thickening (DRT), cystoid macular edema (CME), schitic retinal thickening (SRT) and neurosensory detachment (NSD) of fovea. Some retinal loci presented with a mixture of above patterns There was significant difference in retinal thickness between groups (P<0.001). Focal retinal sensitivity measurement revealed relatively preserved retinal sensitivity in areas with DRT (13.8 dB), moderately reduced sensitivity (7.9 dB) in areas with CME, and gross retinal sensitivity loss in areas with SRT (1.2 dB) and NSD (4.7 dB) (P<0.001). Analysis of regional scotoma depth demonstrated similar pattern. Retinal sensitivity showed better correlation to OCT pattern (r=-0.68, P<0.001) than retinal thickness (r=-0.44, P<0.001). Structure-function correlation allows better understanding of the pathophysiology of visual loss in different morphological types of DME. Classification of macular edema into these categories has implications on the prognosis and predictive value of treatment.

Global levels of H3K27me3 track with differentiation in vivo and are deregulated by MYC in prostate cancer.

Cancer cells and stem cells share a number of biological characteristics including abundant amounts of decondensed chromatin. However, the molecular correlates and the factors involved in altering chromatin structure in cancer cells are not well known. Here, we report that less differentiated stem-like cells in the basal compartment of human and mouse prostate contain lower levels of the polycomb heterochromatin marker H3K27me3 than more differentiated luminal cells. This link to differentiated normal cells is also found in a number of other human and rodent tissues characterized by hierarchical differentiation. In addition to MYC's traditional role as a gene-specific transcription factor, recent studies indicate that MYC also affects global chromatin structure where it is required to maintain "open" or active chromatin. We now demonstrate that in both MYC-driven prostate cancers in mice and human prostate cancers, global levels of H3K27me3 are reduced in prostatic intraepithelial neoplasia and invasive adenocarcinoma lesions. Moreover, decreased levels of H3K27me3 correlate with increased markers of disease aggressiveness (eg, Gleason score and pathological stage). In vitro, experimentally forced reductions in MYC levels result in increased global levels of H3K27me3. These findings suggest that increased levels of decondensed chromatin in both normal progenitor cells and cancer cells are associated with global loss of H3K27me3, which is linked to MYC overexpression.

Differential epithelium DNA damage response to ATM and DNA-PK pathway inhibition in human prostate tissue culture.

The ability of cells to respond and repair DNA damage is fundamental for the maintenance of genomic integrity. Ex vivo culturing of surgery-derived human tissues has provided a significant advancement to assess DNA damage response (DDR) in the context of normal cytoarchitecture in a non-proliferating tissue. Here, we assess the dependency of prostate epithelium DDR on ATM and DNA-PKcs, the major kinases responsible for damage detection and repair by nonhomologous end-joining (NHEJ), respectively. DNA damage was caused by ionizing radiation (IR) and cytotoxic drugs, cultured tissues were treated with ATM and DNA-PK inhibitors, and DDR was assessed by phosphorylation of ATM and its targets H2AX and KAP1, a heterochromatin binding protein. Phosphorylation of H2AX and KAP1 was fast, transient and fully dependent on ATM, but these responses were moderate in luminal cells. In contrast, DNA-PKcs was phosphorylated in both luminal and basal cells, suggesting that DNA-PK-dependent repair was also activated in the luminal cells despite the diminished H2AX and KAP1 responses. These results indicate that prostate epithelial cell types have constitutively dissimilar responses to DNA damage. We correlate the altered damage response to the differential chromatin state of the cells. These findings are relevant in understanding how the epithelium senses and responds to DNA damage.

Global 5-hydroxymethylcytosine content is significantly reduced in tissue stem/progenitor cell compartments and in human cancers.

DNA methylation at the 5-position of cytosines (5 mC) represents an important epigenetic modification involved in tissue differentiation and is frequently altered in cancer. Recent evidence suggests that 5 mC can be converted to 5-hydroxymethylcytosine (5 hmC) in an enzymatic process involving members of the TET protein family. Such 5 hmC modifications are known to be prevalent in DNA of embryonic stem cells and in the brain, but the distribution of 5 hmC in the majority of embryonic and adult tissues has not been rigorously explored. Here, we describe an immunohistochemical detection method for 5 hmC and the application of this technique to study the distribution of 5 hmC in a large set of mouse and human tissues. We found that 5 hmC was abundant in the majority of embryonic and adult tissues. Additionally, the level of 5 hmC closely tracked with the differentiation state of cells in hierarchically organized tissues. The highest 5 hmC levels were observed in terminally differentiated cells, while less differentiated tissue stem/progenitor cell compartments had very low 5 hmC levels. Furthermore, 5 hmC levels were profoundly reduced in carcinoma of the prostate, breast and colon compared to normal tissues. Our findings suggest a distinct role for 5 hmC in tissue differentiation, and provide evidence for its large-scale loss in cancers.

Structural and functional correlates in color vision deficiency.

PURPOSE: The aim of this study is to assess the photoreceptor integrity, using spectral domain optical coherence tomography (SD-OCT), and to measure the retinal sensitivity of patients with congenital red-green color vision deficiency (CVD). METHODS: In all, 14 eyes from 7 patients with congenital red-green CVD (diagnosed by Farnsworth Munsell 100 hue test), and 14 eyes from 7 control subjects were examined by SD-OCT and microperimetry. Radial scans (7-mm) were taken of the macula. The center of the fovea was defined. The thickness of different retinal layers, at the foveal center, and at multiple defined points along all six radial scans, was measured. The median readings were compared between the two groups using Mann-Whitney U-test. RESULTS: SD-OCT demonstrated normal total retinal thickness, normal thickness of the photoreceptor layer, normal thickness of the outer nuclear layer, normal vertical thickness of the outer segments (OSs), and normal vertical thickness of the inner segments. OS horizontal diameter was less in left eye in cases with CVD when compared with controls. The mean retinal and foveal sensitivity was similar between cases and controls. CONCLUSIONS: In subjects with congenital red-green CVD, there are no discernible anatomical abnormalities seen on SD-OCT in various retinal layers, except for a narrower foveal pit. However, further studies with larger sample size are required.

Prevalence of dental caries among 13 and 15-year-old school children in an endemic fluorosis area: a cross-sectional study.

AIM: To assess the prevalence of dental caries and dental fluorosis among 13- to 15-year-old school children in Panyam, Andhra Pradesh, India. MATERIALS AND METHODS: The cross-sectional study was conducted among 202 school children and were examined for dental fluorosis and dental caries. RESULTS: This study shows that male students have a decrease in DMFT (Decayed, missing, filled, teeth) index and increase in Dean's index when compared with females. Among students with 13 to 15 years of age, 13-year-old student has increase in DMFT score when compared with other age groups and 14-year-old students has increase in Dean's score when compared with other age group students. CONCLUSION: The prevalence of dental caries decreased with the increase of fluorosis among the students examined. CLINICAL SIGNIFICANCE: Patients with dental fluorosis show a decreased prevalence of dental caries.

Is neuronal dysfunction an early sign of diabetic retinopathy? Microperimetry and spectral domain optical coherence tomography (SD-OCT) study in individuals with diabetes, but no diabetic retinopathy.

PURPOSE: To elucidate changes in the neurosensory retina in the macular area, using spectral domain OCT and correlate with functional loss on fundus-related microperimetry, in patients with diabetes and no diabetic retinopathy compared with age-matched healthy volunteers. METHODS: This was a prospective study enrolling 39 patients in each group. All patients underwent comprehensive dilated eye examination. The foveal thickness and the photoreceptor layer thickness at the foveal centre were measured using spectral domain OCT, and the mean retinal sensitivity of central 20 degrees was measured using microperimetry. RESULTS: The mean age of the patients with diabetes was 50.92+/-4.75 years, and of controls, 49.87+/-5.50 years. SD-OCT measured photoreceptor layer thickness (PLT) to be 61.62+/-4.48 microm in cases, and 68.79+/-7.84 microm in controls (P<0.0001); foveal thickness (FT) was 168.64+/-16.46 microm in cases and 177.74+/-14.58 microm in controls (P=0.012). The mean retinal sensitivity (MRS) of the central 20 degrees, measured on microperimetry was 15.74+/-3.74 db in cases and 17.70+/-1.5 db in controls (P<0.003). In cases compared with controls (aged under 50 years) statistically significant differences were noted in all the three outcome variables: FT, P=0.030; PLT, P=0.015; and MRS, P=0.020. The duration of diabetes influenced only the PLT (P=0.017). Statistical analysis was performed with Student's t-test and chi2 test. CONCLUSION: Neuronal damage was observed in those eyes that did not have clinical evidence of diabetic retinopathy.

Modelling and optimization of fermentation factors for enhancement of alkaline protease production by isolated Bacillus circulans using feed-forward neural network and genetic algorithm.

AIM: Modelling and optimization of fermentation factors and evaluation for enhanced alkaline protease production by Bacillus circulans. METHODS AND RESULTS: A hybrid system of feed-forward neural network (FFNN) and genetic algorithm (GA) was used to optimize the fermentation conditions to enhance the alkaline protease production by B. circulans. Different microbial metabolism regulating fermentation factors (incubation temperature, medium pH, inoculum level, medium volume, carbon and nitrogen sources) were used to construct a '6-13-1' topology of the FFNN for identifying the nonlinear relationship between fermentation factors and enzyme yield. FFNN predicted values were further optimized for alkaline protease production using GA. The overall mean absolute predictive error and the mean square errors were observed to be 0.0048, 27.9, 0.001128 and 22.45 U ml(-1) for training and testing, respectively. The goodness of the neural network prediction (coefficient of R(2)) was found to be 0.9993. CONCLUSIONS: Four different optimum fermentation conditions revealed maximum enzyme production out of 500 simulated data. Concentration-dependent carbon and nitrogen sources, showed major impact on bacterial metabolism mediated alkaline protease production. Improved enzyme yield could be achieved by this microbial strain in wide nutrient concentration range and each selected factor concentration depends on rest of the factors concentration. The usage of FFNN-GA hybrid methodology has resulted in a significant improvement (>2.5-fold) in the alkaline protease yield. SIGNIFICANCE AND IMPACT OF THE STUDY: The present study helps to optimize enzyme production and its regulation pattern by combinatorial influence of different fermentation factors. Further, the information obtained in this study signifies its importance during scale-up studies.

Production of transgenic rice with agronomically useful genes: an assessment.

Rice is the most important food crop in tropical and subtropical regions of the world. Yield enhancement to increase rice production is one of the essential strategies to meet the demand for food of the growing population. Both abiotic and biotic features limit adversely the productivity of rice growing areas. Conventional breeding has been an effective means for developing high yielding varieties, however; it is associated with its own limitations. It is envisaged that recent trends in biotechnology can contribute to the agronomic improvement of rice in terms of yield and nutritional quality as a supplement to traditional breeding methods. Genetic transformation of rice has demonstrated numerous important opportunities resulting in the genetic improvement of existing elite rice varieties and production of new plant types. Significant advances have been made in the genetic engineering of rice since the first transgenic rice plant production in the late 1980s. Several gene transfer protocols have been employed successfully for the introduction of foreign genes to rice. In more than 60 rice cultivars belonging to indica, japonica, javanica, and elite African cultivars, the protocol has been standardized for transgenic rice production. Selection and use of appropriate promoters, selectable markers, and reporter genes has been helpful for development of efficient protocols for transgenic rice in a number of rice cultivars. The present review is an attempt to assess the current state of development in transgenic rice for the transfer of agronomically useful genes, emphasizing the application and future prospects of transgenic rice production for the genetic improvement of this food crop.

Hypohidrotic Ectodermal Dysplasia with Mitral Valve Prolapse.

Clinical and investigational features of a case of congenital hypohidrotic ectodermal dysplasia in a young boy are described. In addition, he was found to have mitral valve prolapse, which was confirmed by echocardiography. The available literature is discussed. Such an association, though probably fortuitous has not been previously reported.