Exploring the Connection Between Domestic Violence and Masticatory Outcomes in the Pediatric Population: A Systematic Review.

The potential interplay between domestic violence and masticatory outcomes in children and adolescents has garnered increasing attention. Understanding the association between domestic abuse and specific oral health parameters, such as biting habits, temporomandibular disorders (TMDs), and bruxism, holds implications for holistic healthcare interventions. This systematic review aims to synthesize the available evidence to elucidate the potential relationships between domestic abuse and targeted oral health outcomes in the pediatric population. A comprehensive search strategy was conducted across eight databases, namely, PubMed, Embase, Scopus, PsycINFO, Web of Science, Cumulative Index of Nursing and Allied Health Literature (CINAHL), Cochrane Library, and Google Scholar. Boolean operators and Medical Subject Headings (MeSH) keywords were strategically employed to optimize search precision. Clinical studies investigating the relationships between domestic abuse and TMDs, or bruxism, in children and adolescents were included. Two reviewers extracted the data independently. The methodological quality and risk of bias of the selected studies were critically appraised using the Newcastle-Ottawa scale. The systematic search identified three papers investigating the associations between domestic abuse and the targeted oral health parameters. Children in the age group of 6-19 years were assessed. The synthesized evidence revealed a consistent association between domestic abuse and masticatory outcomes. Individuals subjected to domestic abuse exhibited a greater percentage of masticatory anomalies. The methodological assessment of the included studies showed good quality. This systematic review provides a notable synthesis of evidence regarding the associations between domestic abuse and masticatory outcomes in children and adolescents. The complex nature of these relationships necessitates further research to unravel the underlying mechanisms and establish causality. The insights from this review highlight the significance of integrating abuse assessment within oral health evaluations and underscore the need for interdisciplinary collaborations to address the potential impact of abusive experiences on the oral health and well-being of the pediatric population.

Comparison of Continuous and Pulsating Water Jet during Piercing of Ductile Material.

More efficient ways to process materials are constantly being sought, even in the case of continuous water flow technology, which acts on materials mainly by stagnant pressure. An alternative method is an ultrasound-stimulated pulsating water jet, the basis of which is the repeated use of impact pressure, which reduces the time interval for mechanical relaxation. This article focuses on a comparative study from the point of view of water mass flow rate on material penetration and its integrity. Relatively low pressures (p = 20, 30, and 40 MPa) with varying nozzle diameters (d = 0.4 and 0.6 mm) were used to identify the effectiveness of the pulsating water jet. The time exposure of the jet at a fixed place was varied from t = 0.5 to 5 s for each experimental condition. The results showed that with an increase in the pressure and diameter values, the disintegration depth increased. In addition, the surface topography and morphology images showed signs of ductile erosion in the form of erosion pits, upheaved surfaces, and crater formation. The microhardness study showed an increase of 10% subsurface microhardness after the action of the pulsating water jet as compared to the original material.

Design and Analysis of a Low-Cost Electronically Controlled Mobile Ventilator, Incorporating Mechanized AMBU Bag, for Patients during COVID-19 Pandemic.

The outbreak of novel COVID-19 has severely and unprecedentedly affected millions of people across the globe. The painful respiratory distress caused during this disease calls for external assistance to the victims in the form of ventilation. The most common types of artificial ventilating units available at the healthcare facilities and hospitals are exorbitantly expensive to manufacture, and their number is fairly inadequate even in the so-called developed countries to cater to the burning needs of an ever-increasing number of ailing human subjects. According to available reports, without the provision of ventilation, the novel COVID-19 patients are succumbing to their ailments in a huge number of cases. This colossal problem of the availability of ventilator units can be addressed to a great extent by readily producible and cost-effective ventilating units that can be used on those suffering patients during an acute emergency and in the absence of conventional expensive ventilators at hospitals and medical care units. This paper has made an attempt to design and simulate a simple, yet effective, mechanized ventilator unit, which can be conveniently assembled without a profuse skillset and operated to resuscitate an ailing human patient. The stepper motor-controlled kinematic linkage is designed to deliver the patient with a necessitated discharge of air at optimum oxygen saturation through the AMBU bag connected in a ventilation circuit. With the associated code on MATLAB, the motor control parameters such as angular displacement and speed are deduced according to the input patient conditions (age group, tidal volume, breathing rate, etc.) and thereafter fed to the controller that drives the stepper motor. With a proposed feedback loop, the real-time static and dynamic compliance, airway resistance values can be approximately determined from the pressure variation cycle and fed to the controller unit to adjust the tidal volume as and when necessary. The simplistic yet robust design not only renders easy manufacturability by conventional and rapid prototyping techniques like 3D printing at different scales but also makes the product easily portable with minimal handling difficulty. Keeping the motto of Health for All as envisioned by the WHO, this low-cost indigenously engineered ventilator will definitely help the poor and afflicted towards their right to health and will help the medical professionals buy some time to manage the patient with acute respiratory distress syndrome (ARDS) towards recovery. Moreover, this instrument mostly includes readily available functional units having standard specifications and can be considered as standard bought-out items.

Enhancer Dependent Repositioning of TCRb Locus with Respect to the Chromosome Territory.

Intranuclear position of several genes is dynamically altered during development concordant with their activation. To understand this dynamic, but non-random, nuclear organization, it is important to identify the relevant regulatory elements and trans acting factors. Murine TCRb locus gets activated during thymic development. Enhancer Eb is important for VDJ recombination at TCRb locus as it is critically required for establishment of recombination center. Our analysis revealed that TCRb locus gets located out of the chromosome territory specifically in developing thymocytes. Further, CRISPR/Cas9 based deletion mutagenesis established an unambiguous role of enhancer Eb in defining TCRb location relative to chromosome territory. The ability to reposition the target locus relative to chromosome territory highlights a novel aspect pertaining to activity of enhancers which may contribute to their ability to regulate gene expression. Additionally, our observations have implications for understanding the role of enhancers in three-dimensional genome organization and function.

A Two-Stage Process for Differentiation of Wharton's Jelly-Derived Mesenchymal Stem Cells into Neuronal-like Cells.

With no permanent cure for neurodegenerative diseases, the symptoms reappear shortly after the withdrawal of medicines. A better treatment outcome can be expected if the damaged neurons are partly replaced by functional neurons and/or they are repaired using trophic factors. In this regard, safe cell therapy has been considered as a potential alternative to conventional treatment. Here, we have described a two-stage culture process to differentiate Wharton Jelly mesenchymal stem cells (WJ-MSCs) into neuronal-like cells in the presence of various cues involved in neurogenesis. The fate of cells at the end of each stage was analyzed at the morphometric, transcriptional, and translational levels. In the first stage of priming, constitutively, wingless-activated WJ-MSCs crossed the lineage boundary in favor of neuroectodermal lineage, identified by the loss of mesenchymal genes with concomitant expression of neuron-specific markers, like SOX1, PAX6, NTRK1, and NEUROD2. Neuronal-like cells formed in the second stage expressed many mature neuronal proteins like Map2, neurofilament, and Tuj1 and possessed axon hillock-like structures. In conclusion, the differentiation of a large number of neuronal-like cells from nontumorigenic and trophic factors secreting WJ-MSCs promises the development of a therapeutic strategy to treat neurodegenerative diseases.

Characterization of transcripts emanating from enhancer Eß of the murine TCRß locus.

Enhancers are well established as critical regulators of gene expression, but the mechanisms underlying the molecular basis of their specificity and activity are only partly understood. One of the most exciting recent observations is the discovery of enhancer RNA (eRNA), a class of noncoding RNAs derived from enhancer regions. Transcription of developmentally regulated enhancers has been observed to be associated with their active state. The nature of transcripts (eRNA) and their functional attributes are diverse and context dependent. The majority of eRNA are nonpolyadenylated and present in low abundance owing to their low stability, and may represent transcriptional noise. However, some eRNAs have been reported to be reasonably long and stable, are enriched in nuclei, exhibit tissue-specific expression and may contribute to enhancer function. Transcription of enhancers has been postulated to mediate enhancer function through either the act of transcription or via the transcribed RNA per se and is a useful feature to be analysed to understand mechanisms underlying enhancer activity. Enhancer Eß at the murine TCRß locus has been reported to exhibit enhanced occupancy of RNA polymerase II in developing thymocytes. Here, we investigated the transcriptional potential of Eß in developing thymocytes and detected overlapping bidirectional transcripts at Eß ranging between 0.7 and 1.7 kb. These noncoding transcripts are capped, polyadenylated, nuclear and expressed specifically in thymocytes. Delineation of these characteristics is important to further investigate their functional roles in mediating enhancer activity.

Effect of Periodic Water Clusters on AISI 304 Welded Surfaces.

This study compared the effect of the interaction time of periodic water clusters on the surface integrity of AISI 304 tungsten inert gas (TIG) welded joints at different excitation frequencies, as the effect of the technological parameters of pulsating water jet (PWJ) on the mechanical properties of TIG welded joints are under-researched. The TIG welded joints were subjected to different frequencies (20 and 40 kHz) and traverse speeds (1-4 mm/s) at a water pressure of 40 MPa and a standoff distance of 70 mm. The effect of the interaction of the pulsating jet on the material and the enhancement in its mechanical properties were compared through residual stress measurements, surface roughness, and sub-surface microhardness. A maximum enhancement in the residual stress values of up to 480 MPa was observed in the heat-affected zone, along with a maximum roughness of 6.03 µm and a maximum hardness of 551 HV using a frequency of 40 kHz. The improvement in the surface characteristics of the welded joints shows the potential of utilizing pulsed water jet technology with an appropriate selection of process parameters in the treatment of welded structures.

Standoff Distance in Ultrasonic Pulsating Water Jet.

The water hammer effect is the basis of technologies which is artificially responsible for the decay of continuous jets. A recently developed technique enhances the pressure fluctuations using an acoustic chamber, leading to enhanced erosion effects for various water volume flow rates. The optimum standoff distance for an ultrasonic enhanced water jet is not appropriately estimated using an inclined trajectory. The objective of this study is to comprehend the true nature of the interaction of the standoff distance following the stair trajectory and traverse speed of the nozzle on the erosion depth. Additionally, it also critically compares the new method (staircase trajectory) that obeys the variation in frequency of the impingements for defined volume flow rates with the inclined trajectory. In this study, at constant pressure (p = 70 MPa), the role of impingement distribution with the variation of traverse speed (v = 5-35 mm/s) along the centerline of the footprint was investigated. The maximum erosion depth corresponding to each traverse speed is observed at approximately same standoff distance (65 ± 5 mm) and decreases with the increment in traverse speed (h= 1042 and 47 µm at v = 5 and 35 mm/s, respectively). The results are attributed to the variation in the number of impingements per unit length. The surface and morphology analysis of the cross-section using SEM manifested the presence of erosion characteristics (micro-cracks, cavities, voids, and upheaved surface). By varying the water cluster, different impingement densities can be achieved that are suitable for technological operations such as surface peening, material disintegration, or surface roughening.

Coronavirus: An emergency for healthcare professionals.

Coronavirus infection is a transmissible disease. It was first described in China in December, 2019. It has been said to have a person-to-person transmission after prolonged and unprotected exposure. Patients with a potential SARS-CoV-2 exposure present with symptoms of low-grade pyrexia, dry cough, or shortness of breath. People with these symptoms should contact health-care providers before seeking medical intervention so that appropriate preventive actions may be implemented. Health-care facilities should rapidly isolate suspected individuals and notify local health departments for support involved in performing laboratory tests and efforts in containment. The present article describes the nature of virus, method of detection, and its mode of transmission.

Strand-specific detection of overlapping transcripts via purification involving denaturation of biotinylated cDNA.

Reverse transcription-PCR (RT-PCR) is the most widely employed technique for gene expression analysis owing to its high sensitivity, easy reproducibility and fast output. It has been conceived that priming RT reactions with gene-specific primers generates cDNA only from the specific RNA. However, several reports have revealed that cDNA is synthesized even without addition of exogenous primers in RT reactions. Owing to such self-priming activity, the signals from specific strands cannot be accurately detected and can confound the expression analysis, especially in context of overlapping bidirectional transcripts. Here, we demonstrate that purification of biotin-tagged cDNA in conjunction with alkaline denaturation can obviate the problem of background priming and enable accurate strand-specific detection of overlapping transcripts.

An Environmentally Friendly Engineered Azotobacter Strain That Replaces a Substantial Amount of Urea Fertilizer while Sustaining the Same Wheat Yield.

In our endeavor to improve the nitrogen fixation efficiency of a soil diazotroph that would be unaffected by synthetic nitrogenous fertilizers, we have deleted a part of the negative regulatory gene nifL and constitutively expressed the positive regulatory gene nifA in the chromosome of Azotobacter chroococcum CBD15, a strain isolated from the local field soil. No antibiotic resistance gene or other foreign gene was present in the chromosome of the engineered strain. Wheat seeds inoculated with this engineered strain, which we have named Azotobacter chroococcum HKD15, were tested for 3 years in pots and 1 year in the field. The yield of wheat was enhanced by ∼60% due to inoculation of seeds by A. chroococcum HKD15 in the absence of any urea application. Ammonium only marginally affected acetylene reduction by the engineered Azotobacter strain. When urea was also applied, the same wheat yield could be sustained by using seeds inoculated with A. chroococcum HKD15 and using ∼85 kg less urea (∼40 kg less nitrogen) than the usual ∼257 kg urea (∼120 kg nitrogen) per hectare. Wheat plants arising from the seeds inoculated with the engineered Azotobacter strain exhibited far superior overall performance, had much higher dry weight and nitrogen content, and assimilated molecular 15N much better. A nitrogen balance experiment also revealed much higher total nitrogen content. Indole-3-acetic acid (IAA) production by the wild type and that by the engineered strain were about the same. Inoculation of the wheat seeds with A. chroococcum HKD15 did not adversely affect the microbial population in the field rhizosphere soil.IMPORTANCE Application of synthetic nitrogenous fertilizers is a standard agricultural practice to augment crop yield. Plants, however, utilize only a fraction of the applied fertilizers, while the unutilized fertilizers cause grave environmental problems. Wild-type soil diazotrophic microorganisms cannot replace synthetic nitrogenous fertilizers, as these reduce atmospheric nitrogen very inefficiently and almost none at all in the presence of added nitrogenous fertilizers. If the nitrogen-fixing ability of soil diazotrophs could be improved and sustained even in the presence of synthetic nitrogenous fertilizers, then a mixture of the bacteria and a reduced quantity of chemical nitrogenous fertilizers could be employed to obtain the same grain yield but at a much-reduced environmental cost. The engineered Azotobacter strain that we have reported here has considerably enhanced nitrogen fixation and excretion abilities and can replace ∼85 kg of urea per hectare but sustain the same wheat yield, if the seeds are inoculated with it before sowing.

JMJD3 aids in reprogramming of bone marrow progenitor cells to hepatic phenotype through epigenetic activation of hepatic transcription factors.

The strictly regulated unidirectional differentiation program in some somatic stem/progenitor cells has been found to be modified in the ectopic site (tissue) undergoing regeneration. In these cases, the lineage barrier is crossed by either heterotypic cell fusion or direct differentiation. Though studies have shown the role of coordinated genetic and epigenetic mechanisms in cellular development and differentiation, how the lineage fate of adult bone marrow progenitor cells (BMPCs) is reprogrammed during liver regeneration and whether this lineage switch is stably maintained are not clearly understood. In the present study, we wanted to decipher genetic and epigenetic mechanisms that involve in lineage reprogramming of BMPCs into hepatocyte-like cells. Here we report dynamic transcriptional change during cellular reprogramming of BMPCs to hepatocytes and dissect the epigenetic switch mechanism of BM cell-mediated liver regeneration after acute injury. Genome-wide gene expression analysis in BM-derived hepatocytes, isolated after 1 month and 5 months of transplantation, showed induction of hepatic transcriptional program and diminishing of donor signatures over the time. The transcriptional reprogramming of BM-derived cells was found to be the result of enrichment of activating marks (H3K4me3 and H3K9Ac) and loss of repressive marks (H3K27me3 and H3K9me3) at the promoters of hepatic transcription factors (HTFs). Further analyses showed that BMPCs possess bivalent histone marks (H3K4me3 and H3K27me3) at the promoters of crucial HTFs. H3K27 methylation dynamics at the HTFs was antagonistically regulated by EZH2 and JMJD3. Preliminary evidence suggests a role of JMJD3 in removal of H3K27me3 mark from promoters of HTFs, thus activating epigenetically poised hepatic genes in BMPCs prior to partial nuclear reprogramming. The importance of JMJD3 in reprogramming of BMPCs to hepatic phenotype was confirmed by inhibiting catalytic function of the enzyme using small molecule GSK-J4. Our results propose a potential role of JMJD3 in lineage conversion of BM cells into hepatic lineage.

Chromatin Domain Organization of the TCRb Locus and Its Perturbation by Ectopic CTCF Binding.

CTCF-mediated chromatin interactions influence organization and function of mammalian genome in diverse ways. We analyzed the interactions among CTCF binding sites (CBS) at the murine TCRb locus to discern the role of CTCF-mediated interactions in the regulation of transcription and VDJ recombination. Chromosome conformation capture analysis revealed thymocyte-specific long-range intrachromosomal interactions among various CBS across the locus that were relevant for defining the limit of the enhancer Eb-regulated recombination center (RC) and for facilitating the spatial proximity of TCRb variable (V) gene segments to the RC. Ectopic CTCF binding in the RC region, effected via genetic manipulation, altered CBS-directed chromatin loops, interfered with RC establishment, and reduced the spatial proximity of the RC with Trbv segments. Changes in chromatin loop organization by ectopic CTCF binding were relatively modest but influenced transcription and VDJ recombination dramatically. Besides revealing the importance of CTCF-mediated chromatin organization for TCRb regulation, the observed chromatin loops were consistent with the emerging idea that CBS orientations influence chromatin loop organization and underscored the importance of CBS orientations for defining chromatin architecture that supports VDJ recombination. Further, our study suggests that in addition to mediating long-range chromatin interactions, CTCF influences intricate configuration of chromatin loops that govern functional interactions between elements.

Evaluation of Moyer's mixed dentition space analysis in Indian children.

AIM AND OBJECTIVES: Tooth size prediction values are not universal for all ethnic and racial groups. The present study evaluated the applicability of Moyer's mixed dentition space analysis in the Marwari community of Rajasthan, India. MATERIALS AND METHODS: The mesiodistal dimension of permanent mandibular incisors, maxillary and mandibular canine, and premolars of both sides were measured and averaged in 200 adolescents (100 males and 100 females) of the Marwari population in Rajasthan using digital Vernier caliper. Data were statistically analyzed using Student's t-test. The data were then compared with Moyer's predicted values. The tooth measurements of male and female participants were compared with unpaired t-test. RESULTS: Moyer's prediction chart was not comparable with the study population group. The coefficient of correlation and coefficient of determination in our study was 0.57 and 0.25, respectively. Mesiodistal width of measured teeth was lesser in females compared to males for both canine and premolars (P = 0.471 and P = 0.0001, respectively). CONCLUSION: There was significant statistical difference between values of the present study and Moyer's prediction values. Hence, new regression equation and prediction table can be used to predict mesiodistal dimensions of canine and premolars in Marwari children of Rajasthan.

In vitro evaluation of shear bond strength and microleakage of different pit and fissure sealants.

AIM AND OBJECTIVES: Fissure caries is most common in children due to deep pit and fissures. Pit and fissure areas on the occlusal surface of the teeth make them susceptible to dental caries, which need to be prevented or restored. Fissures sealant reduces the risk of occlusal caries. The present study was done to evaluate microleakage and shear bond strength of various fissure sealants. MATERIALS AND METHODS: Thirty-six extracted molars were randomly allocated equally (n = 12) into three groups with three different sealants to evaluate shear bond strength and microleakage at sealant space. The shear bond strengths was evaluated with one-way analysis of variance and microleakage by Kruskal-Wallis nonparametric test using the Statistical Package for the Social Sciences version 18.0 (Chicago: SPSS Inc, 2009). RESULTS: Tetric flow (16.8 MPa) recorded the highest shear bond strength and the difference was statistically significant with enamel loc (12.8 MPa). There was no statistically significant difference in relation to microleakage (P > 0.05) in the tested groups. CONCLUSIONS: Tetric flow recorded the highest shear bond strength and the difference was statistically significant with enamel loc. However, there was no statistically significant difference among the groups regarding microleakage.

Influence of a CTCF-Dependent Insulator on Multiple Aspects of Enhancer-Mediated Chromatin Organization.

Developmental stage-specific enhancer-promoter-insulator interactions regulate the chromatin configuration necessary for transcription at various loci and additionally for VDJ recombination at antigen receptor loci that encode immunoglobulins and T-cell receptors. To investigate these regulatory interactions, we analyzed the epigenetic landscape of the murine T-cell receptor ß (TCRß) locus in the presence and absence of an ectopic CTCF-dependent enhancer-blocking insulator, H19-ICR, in genetically manipulated mice. Our analysis demonstrated the ability of the H19-ICR insulator to restrict several aspects of enhancer-based chromatin alterations that are observed during activation of the TCRß locus for transcription and recombination. The H19-ICR insulator abrogated enhancer-promoter contact-dependent chromatin alterations and additionally prevented Eß-mediated histone modifications that have been suggested to be independent of enhancer-promoter interaction. Observed enhancer-promoter-insulator interactions, in conjunction with the chromatin structure of the Eß-regulated domain at the nucleosomal level, provide useful insights regarding the activity of the regulatory elements in addition to supporting the accessibility hypothesis of VDJ recombination. Analysis of H19-ICR in the heterologous context of the developmentally regulated TCRß locus suggests that different mechanisms proposed for CTCF-dependent insulator action might be manifested simultaneously or selectively depending on the genomic context and the nature of enhancer activity being curtailed.

An ectopic CTCF-dependent transcriptional insulator influences the choice of Vß gene segments for VDJ recombination at TCRß locus.

Insulators regulate transcription as they modulate the interactions between enhancers and promoters by organizing the chromatin into distinct domains. To gain better understanding of the nature of chromatin domains defined by insulators, we analyzed the ability of an insulator to interfere in VDJ recombination, a process that is critically dependent on long-range interactions between diverse types of cis-acting DNA elements. A well-established CTCF-dependent transcriptional insulator, H19 imprint control region (H19-ICR), was inserted in the mouse TCRß locus by genetic manipulation. Analysis of the mutant mice demonstrated that the insulator retains its CTCF and position-dependent enhancer-blocking potential in this heterologous context in vivo. Remarkably, the inserted H19-ICR appears to have the ability to modulate cis-DNA interactions between recombination signal sequence elements of the TCRß locus leading to a dramatically altered usage of Vß segments for Vß-to-DßJß recombination in the mutant mice. This reveals a novel ability of CTCF to govern long range cis-DNA interactions other than enhancer-promoter interactions and suggests that CTCF-dependent insulators may play a diverse and complex role in genome organization beyond transcriptional control. Our functional analysis of mutated TCRß locus supports the emerging role of CTCF in governing VDJ recombination.

Stones from cancerous and benign gallbladders are different: A proton nuclear magnetic resonance spectroscopy study.

AIM: Gallbladder cancer (GBC) is frequently associated with gallstones (GS). At the same time, however, a very small number of patients with GS develop GBC. Cholesterol and metal salts are the common constituents of all GS. To understand their role in the etiopathogenesis of GBC, cholesterol, calcium, and magnesium composition in GS is compared in cancerous and benign gallbladders. METHODS: GS from patients with GBC (n = 11), chronic cholecystitis (CC; n = 23), and xanthogranulomatous cholecystitis (XGC; n = 11) undergoing cholecystectomy were analyzed using proton nuclear magnetic resonance spectroscopy. The diagnosis of the gallbladder disease was based on histopathological examinations. Cholesterol, calcium, and magnesium in the GS of GBC, XGC, and CC were analyzed, compared, and correlated using statistical methods. RESULTS: The quantity of cholesterol was significantly less in the GS of GBC than in benign gallbladder diseases (CC or XGC, P < 0.0001 for both). Both calcium and magnesium were significantly higher in GBC than in benign disease (calcium: P < 0.0005 and magnesium: P < 0.0001 for GBC vs CC; calcium: P < 0.02 and magnesium: P < 0.04 for GBC vs XGC). In all the GS, calcium was higher than magnesium. Calcium and magnesium were positively correlated in GBC (R = 0.69) and XGC (R = 0.75), and cholesterol and calcium were negatively correlated in CC (R =-0.61). CONCLUSION: Differences in the GS composition between malignant and benign gallbladder patients may provide useful clues to the etiopathogenesis of GBC. These clues could lead to the identification of patients with GS in vivo who are at high risk of developing GBC, and advocate prophylactic cholecystectomy to prevent GBC.

Enhancer blocking activity of the insulator at H19-ICR is independent of chromatin barrier establishment.

Transcriptional insulators are cis regulatory elements that organize chromatin into independently regulated domains. At the imprinted murine Igf2/H19 locus, the H19-ICR insulator prevents the activation of the Igf2 promoter on the maternal allele by enhancers that activate H19 on the same chromosome. Given the well-demonstrated role of H19-ICR as an enhancer blocker, we investigated its ability to define a chromatin barrier, as the two activities are coincident on several insulators and may act in concert to define a functional chromatin boundary between adjacent genes with distinct transcriptional profiles. Allele-specific association of posttranslationally modified histones, reflecting the presence of active or inactive chromatin, was analyzed in the region encompassing H19-ICR using chromatin immunoprecipitation. The existence of differential histone modifications upstream and downstream of H19-ICR specifically on the maternal chromosome was observed, which is suggestive of a chromatin barrier formation. However, H19-ICR deletion analysis indicated that distinct chromatin states exist despite the absence of an intervening "barrier." Also, the enhancers can activate the Igf2 promoter despite some parts of the intervening chromatin being in the silent state. Hence, H19-ICR insulator activity is not dependent on preventing the enhancer-mediated alteration of the histone modifications in the region between the Igf2 promoter and the cognate enhancers.

Variation in Different Lipoprotein Fractions if Estimated by Different Methodologies.

BACKGROUND: Determining the levels of lipids and lipoprotein fractions is important in assessing the risk of coronary artery disease. The levels of total cholesterol, triglycerides and high-density lipoproteins are determined directly by enzymatic assays. In the case of low-density lipoproteins and very-low-density lipoproteins, Friedewald's formula has been in use since 1972. According to the formula, the level of very-low-density lipoprotein is equal to that of triglycerides divided by five, while that of low-density lipoprotein cholesterol is equal to the total cholesterol level minus the concentrations of high-density and very-low-density lipoproteins. The determination of low-density lipoprotein by this equation involves three independent lipid analyses, each of which may introduce errors. Besides, other lipoproteins, like intermediate-density lipoproteins and chylomicrons, are not accounted for. This study was done to compare the values of low-density and very-low-density lipoproteins by Friedewald's formula and to determine the values of high-density and low-density lipoproteins by homogenous assays with electrophoretic separation of lipoproteins. METHODS AND RESULTS: Sixty adult patients with triglyceride levels of less than 400 mg/dL were evaluated. The level of low-density lipoprotein was measured enzymatically, using cholesterol esterase, after selective micellary solubilization of low-density lipoprotein cholesterol by a non-ionic detergent. High-density lipoprotein cholesterol was estimated similarly, after binding anti-human ss-lipoprotein antibody to low-density lipoprotein, very-low-density lipoprotein and chylomicrons. Electrophoretic separation of serum lipoproteins was done on agarose gel, and the bands scanned by a densitometer using a 570-nm filter. The values of low-density and very-low-density lipoproteins were also calculated using Friedewald's formula, following the spectrophotometric assay of serum cholesterol and triglycerides. Using the paired t-test, we found that the values of low-density lipoprotein as calculated by Friedewald's formula were significantly different from those determined by direct assays (p < 0.001) and electrophoresis (p < 0.001). Further, the values determined by direct assay and electrophoresis did not differ significantly (p = 0.53). The high-density lipoprotein cholesterol values as determined by direct assay and electrophoresis differed significantly (p < 0.001) from each other. The level of very-low-density lipoprotein as calculated by Friedewald's formula was significantly different from that estimated by electrophoresis (p < 0.001). CONCLUSION: Though Friedewald's equation is widely used in the case of triglyceride levels below 400 mg/dL, the values arrived at are erroneous if there are alterations in intermediate-density lipoproteins, as reported in diabetes mellitus, chronic renal failure, coronary heart disease and certain other disorders. This study shows that even at low triglyceride levels, the calculated values of various lipoproteins differ from those measured by direct spectrophotometric and electrophoretic assays.