Immunohistochemical Expression of Matrix Metalloprotease-2 and Matrix Metalloprotease-9 in the Disks of Patients with Temporomandibular Joint Dysfunction

Purpose Matrix metalloproteases (MMPs) are tissue-remodeling enzymes that function during the remodeling process, such as in immune-inflammatory diseases. Metalloprotease-2 (MMP-2) and metalloprotease-9 (MMP-9) are gelatinases that degrade several types of extracellular matrix collagen. It is hypothesized that in temporomandibular joint (TMJ) dysfunction, MMP-2 and MMP-9 expression levels may be elevated. Therefore, the objective of this study is to determine the association of MMP-2 and MMP-9 expression with temporomandibular joint dysfunction using an immunohistochemical approach to evaluate the joint disk. Material and Methods A total of 45 human temporomandibular joint samples were collected, with 36 samples in the test group (patients with anterior disk displacement with reduction (n = 29) and without reduction (n = 7)) and nine samples in the control group. The immunostaining of the TMJ disks was statistically compared between the groups (P \u3c 0.05). Results There was a statistically significant difference for the area of MMP-2 immunostaining between the control group and the displacement disks with reduction group (ADDwR) (P = 0.048) and between the groups with disk displacement and without reduction (ADDwoR) (P = 0.029). The expression of MMP-2 was significantly elevated in the ADDwoR group. Conclusion No statistically significant difference was found between the variable area of MMP-9 expression in the disk with and without disk displacement, as determined by immunohistochemical analysis. However, there was an elevation of MMP-2 expression in the disks of patients with displacement and without reduction (more severe alteration)

Interleukin-6 Expression in Disc Derangement of Human Temporomandibular Joint and Association with Osteoarthrosis

The inflammatory process is a coordinated response that protects host after infection or trauma, involving several molecular reactions. Once the inflammation is closely linked to the process of destruction of the temporomandibular joint, this study aims to examine, by immunohistochemistry, the expression of interleukin-6 (IL-6), an important inflammatory marker, in temporomandibular articular discs of patients with anterior disc displacement with (ADDwR) and without reduction (ADDwoR) and its association with osteoarthrosis (OA). Thirty-eight (n = 38) articular discs were divided into two cutoffs: 1) analysis 1: 4 control (acute pathology), 17 ADDwR, 17 ADDwoR; and 2) analysis 2: without OA (n = 21) and with OA (n = 17). The area of immunostaining was compared statistically between groups (p \u3c 0.05). In the disc samples, no significant differences were observed between the groups ADDwR and ADDwoR, and with and without OA, in respect to the expression of IL-6 by immunohistochemical examination. Future studies should be conducted with a larger sample size, which could clarify the association of the inflammatory mediator IL-6 with temporomandibular joint dysfunction

Effect of life history on microRNA expression during C. elegans development

Animals have evolved mechanisms to ensure the robustness of developmental outcomes to changing environments. MicroRNA expression may contribute to developmental robustness because microRNAs are key post-transcriptional regulators of developmental gene expression and can affect the expression of multiple target genes. Caenorhabditis elegans provides an excellent model to study developmental responses to environmental conditions. In favorable environments, C. elegans larvae develop rapidly and continuously through four larval stages. In contrast, in unfavorable conditions, larval development may be interrupted at either of two diapause stages: The L1 diapause occurs when embryos hatch in the absence of food, and the dauer diapause occurs after the second larval stage in response to environmental stimuli encountered during the first two larval stages. Dauer larvae are stress resistant and long lived, permitting survival in harsh conditions. When environmental conditions improve, dauer larvae re-enter development, and progress through two post-dauer larval stages to adulthood. Strikingly, all of these life history options (whether continuous or interrupted) involve an identical pattern and sequence of cell division and cell fates. To identify microRNAs with potential functions in buffering development in the context of C. elegans life history options, we used multiplex real-time PCR to assess the expression of 107 microRNAs throughout development in both continuous and interrupted life histories. We identified 17 microRNAs whose developmental profile of expression is affected by dauer life history and/or L1 diapause, compared to continuous development. Hence these microRNAs could function to regulate gene expression programs appropriate for different life history options in the developing worm

The \u3cem\u3elet-7\u3c/em\u3e MicroRNA Family Members \u3cem\u3emir\u3c/em\u3e-48, \u3cem\u3emir\u3c/em\u3e-84, and mir-241 Function Together to Regulate Developmental Timing in \u3cem\u3eCaenorhabditis elegans\u3c/em\u3e

The microRNA let-7 is a critical regulator of developmental timing events at the larval-to-adult transition in C. elegans. Recently, microRNAs with sequence similarity to let-7 have been identified. We find that doubly mutant animals lacking the let-7 family microRNA genes mir-48 and mir-84 exhibit retarded molting behavior and retarded adult gene expression in the hypodermis. Triply mutant animals lacking mir-48, mir-84, and mir-241 exhibit repetition of L2-stage events in addition to retarded adult-stage events. mir-48, mir-84, and mir-241 function together to control the L2-to-L3 transition, likely by base pairing to complementary sites in the hbl-1 3′ UTR and downregulating hbl-1 activity. Genetic analysis indicates that mir-48, mir-84, and mir-241 specify the timing of the L2-to-L3 transition in parallel to the heterochronic genes lin-28 and lin-46. These results indicate that let-7 family microRNAs function in combination to affect both early and late developmental timing decisions

Most \u3cem\u3eCaenorhabditis elegans\u3c/em\u3e MicroRNAs are Individually Not Essential for Development or Viability

MicroRNAs (miRNAs), a large class of short noncoding RNAs found in many plants and animals, often act to post-transcriptionally inhibit gene expression. We report the generation of deletion mutations in 87 miRNA genes in Caenorhabditis elegans, expanding the number of mutated miRNA genes to 95, or 83% of known C. elegans miRNAs. We find that the majority of miRNAs are not essential for the viability or development of C. elegans, and mutations in most miRNA genes do not result in grossly abnormal phenotypes. These observations are consistent with the hypothesis that there is significant functional redundancy among miRNAs or among gene pathways regulated by miRNAs. This study represents the first comprehensive genetic analysis of miRNA function in any organism and provides a unique, permanent resource for the systematic study of miRNAs

FasL Expression in Articular Discs of Human Temporomandibular Joint and Association with Osteoarthrosis

Background Apoptosis is a programme of cell death which does not induce an inflammatory response. Recent previous research has suggested a correlation between temporomandibular internal derangement and apoptosis. Fas ligand (FasL) is an apoptosis‐inducing factor, known to trigger apoptosis through distinct signal pathways. This study aims to examine, by immunohistochemistry, the expression of FasL in temporomandibular joint (TMJ) articular discs of patients with anterior disc displacement with reduction (ADDwR) and without reduction (ADDwoR) in patients with and without osteoarthrosis (OA). Methods Forty‐two (n = 42) TMJ articular discs were divided into two cut‐offs: (i) 8 control, 17 ADDwR, 17 ADDwoR, and (ii) without OA (n = 25) and with OA (n = 17). The area of immunostaining was compared statistically between groups (P \u3c 0.05). Results Statistically significant differences were found in the expression of FasL in TMJ discs between the three groups (P = 0.001). ADDwR presented significant higher FasL expression when compared with ADDwoR (P \u3c 0.001). Significant higher FasL expression was observed in the group without OA (P = 0.001). All patients without OA presented ADDwR, while all the patients with OA presented ADDwoR. Conclusion A higher area of in situ immunostaining of FasL was found in temporomandibular discs with reduction, which is the less severe condition. Moreover, a reduced expression of FasL in the discs of patients with osteoarthrosis was found, suggesting that some aspects of apoptosis might underlie the progression of TMJ disorders

Mutations in Conserved Residues of the C. elegans microRNA Argonaute ALG-1 Identify Separable Functions in ALG-1 miRISC Loading and Target Repression

microRNAs function in diverse developmental and physiological processes by regulating target gene expression at the post-transcriptional level. ALG-1 is one of two Caenorhabditis elegans Argonautes (ALG-1 and ALG-2) that together are essential for microRNA biogenesis and function. Here, we report the identification of novel antimorphic (anti) alleles of ALG-1 as suppressors of lin-28(lf) precocious developmental phenotypes. The alg-1(anti) mutations broadly impair the function of many microRNAs and cause dosage-dependent phenotypes that are more severe than the complete loss of ALG-1. ALG-1(anti) mutant proteins are competent for promoting Dicer cleavage of microRNA precursors and for associating with and stabilizing microRNAs. However, our results suggest that ALG-1(anti) proteins may sequester microRNAs in immature and functionally deficient microRNA Induced Silencing Complexes (miRISCs), and hence compete with ALG-2 for access to functional microRNAs. Immunoprecipitation experiments show that ALG-1(anti) proteins display an increased association with Dicer and a decreased association with AIN-1/GW182. These findings suggest that alg-1(anti) mutations impair the ability of ALG-1 miRISC to execute a transition from Dicer-associated microRNA processing to AIN-1/GW182 associated effector function, and indicate an active role for ALG/Argonaute in mediating this transition

Direct visualization of the native structure of viroid RNAs at single-molecule resolution by atomic force microscopy

[EN] Viroids are small infectious, non-protein-coding circular RNAs that replicate independently and, in some cases, incite diseases in plants. They are classified into two families: Pospiviroidae, composed of species that have a central conserved region (CCR) and replicate in the cell nucleus, and Avsunviroidae, containing species that lack a CCR and whose multimeric replicative intermediates of either polarity generated in plastids self-cleave through hammerhead ribozymes. The compact, rod-like or branched, secondary structures of viroid RNAs have been predicted by RNA folding algorithms and further examined using different in vitro and in vivo experimental techniques. However, direct data about their native tertiary structure remain scarce. Here we have applied atomic force microscopy (AFM) to image at single-molecule resolution different variant RNAs of three representative viroids: potato spindle tuber viroid (PSTVd, family Pospiviroidae), peach latent mosaic viroid and eggplant latent viroid (PLMVd and ELVd, family Avsunviroidae). Our results provide a direct visualization of their native, three-dimensional conformations at 0 and 4 mM Mg2+ and highlight the role that some elements of tertiary structure play in their stabilization. The AFM images show that addition of 4 mM Mg2+ to the folding buffer results in a size contraction in PSTVd and ELVd, as well as in PLMVd when the kissing-loop interaction that stabilizes its 3D structure is preserved.This work was supported by the Spanish Ministerio de Economia y Competitividad (MINECO) grants BIO2016-79618-R (funded by EU under the FEDER programme) to C.B. and BFU2104-56812-P to R.F., as well as by the Comunidad de Madrid grant S2018/NMT-4349 to L.V. 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The C. Elegans Heterochronic Gene lin-46 Affects Developmental Timing at Two Larval Stages and Encodes a Relative of the Scaffolding Protein Gephyrin

The succession of developmental events in the C. elegans larva is governed by the heterochronic genes. When mutated, these genes cause either precocious or retarded developmental phenotypes, in which stage-specific patterns of cell division and differentiation are either skipped or reiterated, respectively. We identified a new heterochronic gene, lin-46, from mutations that suppress the precocious phenotypes caused by mutations in the heterochronic genes lin-14 and lin-28. lin-46 mutants on their own display retarded phenotypes in which cell division patterns are reiterated and differentiation is prevented in certain cell lineages. Our analysis indicates that lin-46 acts at a step immediately downstream of lin-28, affecting both the regulation of the heterochronic gene pathway and execution of stage-specific developmental events at two stages: the third larval stage and adult. We also show that lin-46 is required prior to the third stage for normal adult cell fates, suggesting that it acts once to control fates at both stages, and that it affects adult fates through the let-7 branch of the heterochronic pathway. Interestingly, lin-46 encodes a protein homologous to MoeA of bacteria and the C-terminal domain of mammalian gephyrin, a multifunctional scaffolding protein. Our findings suggest that the LIN-46 protein acts as a scaffold for a multiprotein assembly that controls developmental timing, and expand the known roles of gephyrin-related proteins to development

Synthesis of a mitochondria-targeted spin trap using a novel Parham-type cyclization

A new cyclic nitrone spin trap, [4-(3′,3′-dibutyl-2′-oxy-3′H-isoindol-5′-yloxy)butyl]triphenylphosphonium bromide (MitoSpin), bearing a lipophilic cation has been prepared by a route that involves a novel Parham-type lithiation–cyclization of an isocyanate to give the isoindolinone core. MitoSpin accumulates in a membrane potential dependent way in energized mitochondria and its oxidation could potentially be used in the study of oxidative stress resulting from reactive oxygen species generated in mitochondria