Use of Endodontic Procedures in Young Permanent Teeth.

Background: Regenerative endodontic procedures (REPs) have emerged as a promising treatment option for young permanent teeth with pulp necrosis, offering the potential for tissue repair and preservation. Materials and Methods: A retrospective analysis was conducted on a cohort of 30 patients aged 8 to 16 years with pulp necrosis in young permanent teeth. The patients underwent REPs, including disinfection, triple antibiotic paste application, and a coronal barrier. Clinical and radiographic data were collected at baseline and follow-up appointments at 6, 12, and 24 months. Radiographs were analyzed for root lengthening, apical closure, and resolution of periapical lesions. Results: The mean increase in root length after 24 months was 3.42 mm (SD ± 1.12 mm), and 90% of cases demonstrated complete apical closure. The overall success rate, defined as the absence of clinical symptoms and radiographic evidence of pathology, was 80. Conclusion: REPs show promising outcomes in young permanent teeth with pulp necrosis, promoting root development, and apical closure.

P66Shc Mediates SUMO2-induced Endothelial Dysfunction.

Sumoylation is a post-translational modification that can regulate different physiological functions. Increased sumoylation, specifically conjugation of SUMO2/3 (small ubiquitin like modifier 2/3), is detrimental to vascular health. However, the molecular mechanism mediating this effect is poorly understood. Here, we demonstrate that SUMO2 modifies p66Shc, which impairs endothelial function. Using multiple approaches, we show that p66Shc is a direct target of SUMO2. Mass spectrometry identified that SUMO2 modified lysine-81 in the unique collagen homology-2 domain of p66Shc. SUMO2ylation of p66Shc increased phosphorylation at serine-36, causing it to translocate to the mitochondria. Notably, sumoylation-deficient p66Shc (p66ShcK81R) was resistant to SUMO2-induced p66ShcS36 phosphorylation and mitochondrial translocation. Ingenuity pathway analysis showed that majority of effects of p66Shc SUMO2ylation were mediated via p66ShcK81. Finally, p66ShcK81R knockin mice were resistant to SUMO2-induced endothelial dysfunction. Collectively, our work uncovers a posttranslational modification of redox protein p66Shc and identifies SUMO2-p66Shc signaling as a regulator of vascular endothelial function.

Influence of pretreatment exposure to pediatric dental care using the "Tiny dentist" game on 4-7 years old children's pain and anxiety: a parallel randomised clinical trial.

There have been no experiments on interactive modelling through computer games, although there have been a few on modelling a pre-exposure method for managing anxiety among preschoolers. The impact of partaking in a dental simulation game prior to the dental treatment on pain and anxiety in kids aged 4 to7 years during their first appointment was studied. A total of 156 kids who required unilateral pulp therapy and preformed crowns on their mandibular primary molars were enrolled in this double-blind, randomized clinical trial. They were then randomly assigned to intervention and control groups. The intervention group engaged in the game three times/day for 07 days prior to the anticipated appointment. The Wong-Baker Faces Rating Scale (WBFRS) was used to record their pre- and post-operative pain experienced during the dental procedure. Additionally, a finger pulse oximeter was used to record heart rate (HR) at each of the six treatment phases: baseline (the first session, two weeks prior to treatment) and stages 2-6. Playing video games considerably lowered the heart rate. Playing and the treatment period interacted in a major way. On comparing the groups at every time point, the intervention group displayed lower HR during injection, tooth preparation with an air-rotor and biomechanical preparation with endodontic rotary files.The results suggest that engaging in specific dental simulation games prior to the 1st dentist visit could help preschoolers feel less anxious during routine dental operations.

Antibody response in elderly vaccinated four times with an mRNA anti-COVID-19 vaccine.

The humoral response after the fourth dose of a mRNA vaccine against COVID-19 has not been adequately described in elderly recipients, particularly those not exposed previously to SARS-CoV-2. Serum anti-RBD IgG levels (Abbott SARS-CoV-2 IgG II Quant assay) and neutralizing capacities (spike SARS-CoV-2 pseudovirus Wuhan and Omicron BA.1 variant) were measured after the third and fourth doses of a COVID-19 mRNA vaccine among 46 elderly residents (median age 85 years [IQR 81; 89]) of an assisted living facility. Among participants never infected by SARS-CoV-2, the mean serum IgG levels against RBD (2025 BAU/ml), 99 days after the fourth vaccine, was as high as 76 days after the third vaccine (1987 BAU/ml), and significantly higher (p = 0.030) when the latter were corrected for elapsed time. Neutralizing antibody levels against the historical Wuhan strain were significantly higher (Mean 1046 vs 1573; p = 0.002) and broader (against Omicron) (Mean 170 vs 375; p = 0.018), following the fourth vaccine. The six individuals with an Omicron breakthrough infection mounted strong immune responses for anti-RBD and neutralizing antibodies against the Omicron variant indicating that the fourth vaccine dose did not prevent a specific adaptation of the immune response. These findings point out the value of continued vaccine boosting in the elderly population.

Using the Caries Assessment Spectrum and Treatment Index to Determine the Caries Incidence in Primary and Permanent Molars Among School Children of Kolkata: A Cross-Sectional Study.

INTRODUCTION: Caries in their early stages cannot be properly classified using the decayed, missing, and filled teeth (DMFT) index because it does not record precavitated lesions. Thus, the caries assessment spectrum and treatment (CAST) index is a good alternative as it is equipped to record the whole range of dental illnesses and count restored teeth as healthy ones. MATERIALS AND METHODS: The participants in this cross-sectional study included 300 children who were seven-to-eight years of age. We used the CAST index to assess the extent of caries in the deciduous and permanent molars of these children. For this, all the permanent and baby teeth, the primary and secondary permanent molars, as well as the first and second deciduous molars were examined to determine the prevalence of each carious stage. The correlation of the distribution of the CAST codes between the first and second molars, the second and first molars, the right and left sides of the dental bend, and the opposing jaws was analyzed using Spearman's rank correlation coefficient, while the cut-off for statistical significance was a p-value of 0.05. The quantitative analysis was conducted using IBM SPSS Experiences Version 20 for Windows. RESULTS: By comparing CAST codes in the right and left molars, we were able to observe how the development of caries in paired teeth might affect one another. The rank correlation value was found to be less than 0.5 only in the primary second molars (55/65 and 85/75), which were found exclusively in the deciduous first molars. Moreover, the r values for the neighboring deciduous and permanent molars were found to be below 0.3, i.e., 65/64 (0.497), 74/75 (0.327), and 84/85 (0.411), which indicated a weak connection between them. When comparing the teeth in different jaws, we found reasonable correlations (r = 0.33-0.49), with only 64/74 outliers (0.501). CONCLUSIONS: We found that, in the examined population, there was a well-established correlation between the stages of caries development in the deciduous molars on the left side of the mouth.

Indexing Resilience to Heat and Drought Stress in the Wild Relatives of Rapeseed-Mustard.

Wild species are weedy relatives and progenitors of cultivated crops, usually maintained in their centres of origin. They are rich sources of diversity as they possess many agriculturally important traits. In this study, we analysed 25 wild species and 5 U triangle species of Brassica for their potential tolerance against heat and drought stress during germination and in order to examine the early seedling stage. We identified the germplasms based on the mean membership function value (MFV), which was calculated from the tolerance index of shoot length, root length, and biochemical analysis. The study revealed that B. napus (GSC-6) could withstand high temperatures and drought. Other genotypes that were tolerant to the impact of heat stress were B. tournefortii (RBT 2002), D. gomez-campoi, B. tournefortii (Rawa), L. sativum, and B. carinata (PC-6). C. sativa resisted drought but did not perform well when subjected to high temperatures. Tolerance to drought was observed in B. fruticulosa (Spain), B. tournefortii (RBT 2003), C. bursa-pastoris (late), D. muralis, C. abyssinica (EC694145), C. abyssinica (EC400058) and B. juncea (Pusa Jaikisan). This investigation contributes to germplasm characterization and the identification of the potential source of abiotic stress tolerance in the Brassica breeding programme. These identified genotypes can be potential sources for transferring the gene(s)/genomic regions that determine tolerance to the elite cultivars.

Age Estimation by Dental Calcification Stages and Hand-Wrist Radiograph.

INTRODUCTION: An essential part of pediatric dentistry in recent times is age estimation for various purposes such as orthodontics, forensic dentistry, human anthropology, and bioarchaeology. Assessment of calcification of dental tissue is another physiologic method for skeletal growth assessment. AIMS: This study aims to evaluate the correlation between dental calcification stages and skeletal maturity indicators and their application in age estimation purposes. METHODS: Tooth calcification was assessed by Demirjian's method and hand-wrist assessment was done by Fishman's method. Spearman's rank-order correlation coefficient was applied to measure the association between skeletal maturational indicators and dental calcification stages of individual teeth, and the statistical significance of the correlation was tested. RESULTS: Spearman's significant coefficients for canine, first premolar, second premolar, and molar are 0.11, 0.09, 0.09, and 0.13, respectively, which are not significant. CONCLUSION: Fishman's method of hand-wrist radiograph assessment is quite accurate as a maturity indicator but its association with dental calcification stages cannot be established.

Overexpression of UBA5 in Cells Mimics the Phenotype of Cells Lacking UBA5.

Ufmylation is a posttranslational modification in which the modifier UFM1 is attached to target proteins. This conjugation requires the concerted work of three enzymes named UBA5, UFC1, and UFL1. Initially, UBA5 activates UFM1 in a process that ends with UFM1 attached to UBA5's active site Cys. Then, in a trans-thiolation reaction, UFM1 is transferred from UBA5 to UFC1, forming a thioester bond with the latter. Finally, with the help of UFL1, UFM1 is transferred to the final destination-a lysine residue on a target protein. Therefore, not surprisingly, deletion of one of these enzymes abrogates the conjugation process. However, how overexpression of these enzymes affects this process is not yet clear. Here we found, unexpectedly, that overexpression of UBA5, but not UFC1, damages the ability of cells to migrate, in a similar way to cells lacking UBA5 or UFC1. At the mechanistic level, we found that overexpression of UBA5 reverses the trans-thiolation reaction, thereby leading to a back transfer of UFM1 from UFC1 to UBA5. This, as seen in cells lacking UBA5, reduces the level of charged UFC1 and therefore harms the conjugation process. In contrast, co-expression of UBA5 with UFM1 abolishes this effect, suggesting that the reverse transfer of UFM1 from UFC1 to UBA5 depends on the level of free UFM1. Overall, our results propose that the cellular expression level of the UFM1 conjugation enzymes has to be tightly regulated to ensure the proper directionality of UFM1 transfer.

Phosphorylation-dependent association of human chromatin protein PC4 to linker histone H1 regulates genome organization and transcription.

Human Positive Coactivator 4 (PC4) is a multifaceted chromatin protein involved in diverse cellular processes including genome organization, transcription regulation, replication, DNA repair and autophagy. PC4 exists as a phospho-protein in cells which impinges on its acetylation by p300 and thereby affects its transcriptional co-activator functions via double-stranded DNA binding. Despite the inhibitory effects, the abundance of phosphorylated PC4 in cells intrigued us to investigate its role in chromatin functions in a basal state of the cell. We found that casein kinase-II (CKII)-mediated phosphorylation of PC4 is critical for its interaction with linker histone H1. By employing analytical ultracentrifugation and electron microscopy imaging of in vitro reconstituted nucleosomal array, we observed that phospho-mimic (PM) PC4 displays a superior chromatin condensation potential in conjunction with linker histone H1. ATAC-sequencing further unveiled the role of PC4 phosphorylation to be critical in inducing chromatin compaction of a wide array of coding and non-coding genes in vivo. Concordantly, phospho-PC4 mediated changes in chromatin accessibility led to gene repression and affected global histone modifications. We propose that the abundance of PC4 in its phosphorylated state contributes to genome compaction contrary to its co-activator function in driving several cellular processes like gene transcription and autophagy.

CpPosNeg: A positive-negative selection strategy allowing multiple cycles of marker-free engineering of the Chlamydomonas plastome.

The chloroplast represents an attractive compartment for light-driven biosynthesis of recombinant products, and advanced synthetic biology tools are available for engineering the chloroplast genome ( = plastome) of several algal and plant species. However, producing commercial lines will likely require several plastome manipulations. This presents issues with respect to selectable markers, since there are a limited number available, they can be used only once in a serial engineering strategy, and it is undesirable to retain marker genes for antibiotic resistance in the final transplastome. To address these problems, we have designed a rapid iterative selection system, known as CpPosNeg, for the green microalga Chlamydomonas reinhardtii that allows creation of marker-free transformants starting from wild-type strains. The system employs a dual marker encoding a fusion protein of E. coli aminoglycoside adenyltransferase (AadA: conferring spectinomycin resistance) and a variant of E. coli cytosine deaminase (CodA: conferring sensitivity to 5-fluorocytosine). Initial selection on spectinomycin allows stable transformants to be established and driven to homoplasmy. Subsequent selection on 5-fluorocytosine results in rapid loss of the dual marker through intramolecular recombination between the 3'UTR of the marker and the 3'UTR of the introduced transgene. We demonstrate the versatility of the CpPosNeg system by serial introduction of reporter genes into the plastome.

Intramuscular mRNA BNT162b2 vaccine against SARS-CoV-2 induces neutralizing salivary IgA.

Intramuscularly administered vaccines stimulate robust serum neutralizing antibodies, yet they are often less competent in eliciting sustainable "sterilizing immunity" at the mucosal level. Our study uncovers a strong temporary neutralizing mucosal component of immunity, emanating from intramuscular administration of an mRNA vaccine. We show that saliva of BNT162b2 vaccinees contains temporary IgA targeting the receptor-binding domain (RBD) of severe acute respiratory syndrome coronavirus-2 spike protein and demonstrate that these IgAs mediate neutralization. RBD-targeting IgAs were found to associate with the secretory component, indicating their bona fide transcytotic origin and their polymeric multivalent nature. The mechanistic understanding of the high neutralizing activity provided by mucosal IgA, acting at the first line of defense, will advance vaccination design and surveillance principles and may point to novel treatment approaches and new routes of vaccine administration and boosting.

Isolation and optimization of a novel thraustochytrid strain for DHA rich and astaxanthin comprising biomass as aquafeed supplement.

A marine organism, belonging to the Thraustochytrids family was isolated from mangroves of Mumbai, India. The isolated strain was identified as Aurantiochytrium limacinum by internal transcribed spacer sequence analysis. Optimization of process parameters yielded 14.47 g/L dry cell weight containing 55-58% oil in 3.5 days' cultivation on glucose, yeast extract, and peptone in the bioreactor. Docosahexaenoic acid (DHA) was found to be the dominant long-chain polyunsaturated fatty acid, accounting for 32-35% of total fatty acid content. The process parameter was tweaked to simultaneously synthesize astaxanthin along with DHA. The concurrent synthesis of DHA and astaxanthin-containing biomass establishes the isolated strain as a perfect choice for aquafeed. Accession number: NCBI accession number MN046792.

Time-dependent study of graphene oxide-trypsin adsorption interface and visualization of nano-protein corona.

Understanding of interactions of nanomaterials with biomolecules (especially proteins) is of great importance to the area of nanobiotechnology. Graphene and its derivative such as graphene oxide (GO), are two-dimensional (2-D) nanomaterials with remarkable physical and chemical properties and have been broadly explored in biotechnology and biomedical application. Here, we have reported the nature of adsorption of trypsin on the GO surface, considering its biomedical implications. A simple incubation of trypsin on GO surface exhibits varying resistance to autolysis. The structural morphology of trypsin on the GO surface was studied by using atomic force microscopy (AFM), circular dichroism (CD), fluorescence, and total internal reflection fluorescence (TIRF) microscopies. Results suggest that the trypsin follows the Freundlich Isotherm. By the Langmuir model, the maximum adsorption capacity was found to be 100 mg/g. From protein assay results we have concluded that the native trypsin exhibits the highest catalytic efficiency (33.97*104 L mol-1 min-1) in comparison to other Trp-GO constructs. We have further visualized morphological change on GO-trypsin interface throughout the adsorption process by taking samples at definite time intervals, which suggests that the interaction of trypsin with GO is an example of the soft corona. Our findings may be implicated in enzyme engineering as well as enzyme-based bio-sensing applications.

A Novel Synthesis of the Graphene Oxide-Silver (GO-Ag) Nanocomposite for Unique Physiochemical Applications.

Graphene oxide-silver nanocomposite (GO-Ag) was fabricated via the sonochemical method, which shows unique physiochemical properties. Graphene oxide (GO) and silver nanoparticles (AgNPs) were synthesized by modified Hummer's and Chemical reduction methods, respectively. The synthesized nanocomposite was characterized using powder X-ray diffraction, Raman spectroscopy, and Fourier-transform infrared spectroscopy. The surface morphology of synthesized nanoparticles was studied using scanning electron microscopy and transmission electron microscopy. The thermoluminescence property of the nanocomposite was analyzed by irradiating the samples in gamma radiation at 1 kGy. Electrochemical reversibility of the GO-Ag nanocomposite was examined by cyclic voltammetry. The photocatalytic application of the nanocomposite was studied using degradation of methylene blue dye. Results reveal that doping of AgNPs on the GO surface not only improves its dye degradation property but also enhances its thermoluminescence property. This knowledge will be helpful in determining the antibacterial property of the GO-Ag nanocomposite in the future.

ClpB is an essential stress regulator of Mycobacterium tuberculosis and endows survival advantage to dormant bacilli.

The ability to tolerate multiple host derived stresses, resist eradication and persist within the infected individuals is central to the pathogenicity of Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB). Mycobacterial survival is contingent upon sensing environmental perturbations and initiating a fitting response to counter them. Therefore, understanding of molecular mechanisms underlying stress tolerance and sensing in Mtb is critical for devising strategies for TB control. Our study aims to delineate the role of ClpB, a heat shock protein of Hsp100 family, in the general stress response and persistence mechanisms of Mtb. We demonstrate that Mtb requires ClpB to survive under stressful conditions. Additionally, we show that ClpB is necessary for the bacteria to persist in latency-like conditions such as prolonged hypoxia and nutrient-starvation. The disruption of ClpB results in aberrant cellular morphology, impaired biofilm formation and reduced infectivity of Mtb ex vivo. Our study also reports an alternative role of ClpB as a chaperokine which elicits inflammatory response in host. We conclude that ClpB is essential for Mtb to survive within macrophages, and plays a crucial part in the maintenance of dormant Mtb bacilli in latent state. The absence of ClpB in human genome makes it an attractive choice as drug target for TB.

In vitro and in vivo characterization of the interaction, proinflammatory, immunomodulatory and antigenic properties of capsular polysaccharide from Streptococcus pneumoniae serotype 1.

Pneumococcal capsular polysaccharide (PCP) is the major virulence determinant of Streptococcus pneumoniae (pneumococcus). Strains devoid of the capsule are avirulent or highly attenuated. PCP is present in soluble form and on pneumococci in infected individuals. The present study was undertaken to study the interaction of PCP from serotype 1 (PCP1) with immune cells, and its proinflammatory, immunomodulatory and antigenic properties. Binding of PCP1 to the surface of immune cells led to proinflammatory cytokine production which was not cell line or cytokine restricted. HEK293T transfectants expressing TLR1 and TLR2 produced IL-8 upon stimulation with PCP1, untransfected cells did not do so. PCP1 failed to induce TNF-α production from RAW264.7 cells when pre-incubated with a TLR2 blocking antibody. The surface binding of PCP1 was abrogated in the presence of TLR2 blocking antibody. PCP1 failed to bind TLR2 deficient RAW264.7 cells and induce TNF-α production. Unlike PCP1, alkali-treated PCP1 failed to stimulate RAW264.7 cells to produce TNF-α indicating the importance of alkali-sensitive moieties like O-acetyl groups. Alkali-treated PCP1 elicited lower anti-PCP1 antibody response. Mice experiments suggested that alkali-sensitive groups are significant target of protective antibodies in PCP1 immunized mice. Our findings demonstrate that PCP1 is an important modulator of immune response against pneumococci.

Chromatin protein PC4 is downregulated in breast cancer to promote disease progression: Implications of miR-29a.

The human transcriptional coactivator PC4 has numerous roles to play in the cell. Other than its transcriptional coactivation function, it facilitates chromatin organization, DNA damage repair, viral DNA replication, etc. Although it was found to be an essential protein in vivo, the importance of this multifunctional protein in the regulation of different cellular pathways has not been investigated in details, particularly in oncogenesis. In this study, PC4 downregulation was observed in a significant proportion of mammary tissues obtained from Breast cancer patient samples as well as in a subset of highly invasive and metastatic Breast cancer patient-derived cell lines. We have identified a miRNA, miR-29a which potentially reduce the expression of PC4 both in RNA and protein level. This miR-29a was found to be indeed overexpressed in a substantial number of Breast cancer patient samples and cell lines as well, suggesting one of the key mechanisms of PC4 downregulation. Stable Knockdown of PC4 in MCF7 cells induced its migratory as well as invasive properties. Furthermore, in an orthotopic breast cancer mice model system; we have shown that reduced expression of PC4 enhances the tumorigenic potential substantially. Absence of PC4 led to the upregulation of several genes involved in Epithelial to Mesenchymal Transition (EMT), indicating the possible mechanism of uniform tumour progression in the orthotropic mice. Collectively these data establish the role of PC4 in tumour suppression.

Nonhistone human chromatin protein PC4 is critical for genomic integrity and negatively regulates autophagy.

Multifunctional human transcriptional positive co-activator 4 (PC4) is a bona fide nonhistone component of the chromatin and plays a pivotal role in the process of chromatin compaction and functional genome organization. Knockdown of PC4 expression causes a drastic decompaction which leads to open conformation of the chromatin, and thereby altered nuclear architecture, defects in chromosome segregation and changed epigenetic landscape. Interestingly, these defects do not induce cellular death but result in enhanced cellular proliferation, possibly through enhanced autophagic activity. Moreover, PC4 depletion confers significant resistance to gamma irradiation. Exposure to gamma irradiation further induced autophagy in these cells. Inhibition of autophagy by small molecule inhibitors as well as by silencing of a critical autophagy gene drastically reduces the ability of PC4 knockdown cells to survive. On the contrary, complementation with wild-type PC4 could reverse this phenomenon, confirming the process of autophagy as the key mechanism for radiation resistance in the absence of PC4. These data connect the unexplored role of chromatin architecture in regulating autophagy during stress conditions such as radiation.

Loss of Hepatic Oscillatory Fed microRNAs Abrogates Refed Transition and Causes Liver Dysfunctions.

Inability to mediate fed-fast transitions in the liver is known to cause metabolic dysfunctions and diseases. Intuitively, a failure to inhibit futile translation of state-specific transcripts during fed-fast cycles would abrogate dynamic physiological transitions. Here, we have discovered hepatic fed microRNAs that target fasting-induced genes and are essential for a refed transition. Our findings highlight the role of these fed microRNAs in orchestrating system-level control over liver physiology and whole-body energetics. By targeting SIRT1, PGC1α, and their downstream genes, fed microRNAs regulate metabolic and mitochondrial pathways. MicroRNA expression, processing, and RISC loading oscillate during these cycles and possibly constitute an anticipatory mechanism. Fed-microRNA oscillations are deregulated during aging. Scavenging of hepatic fed microRNAs causes uncontrolled gluconeogenesis and failure in the catabolic-to-anabolic switching upon feeding, which are hallmarks of metabolic diseases. Besides identifying mechanisms that enable efficient physiological toggling, our study highlights fed microRNAs as candidate therapeutic targets.