bluuebunny/crossref_metadata_2025_split huggingface dataset:
Bifidobacterium dentium Modulates Expression of Genes Important to Synaptic Plasticity and Neurodevelopment in the Cerebellum of Germ‐Free Mice
Authors: Luk Berkley , Engevik Melinda , Hall Anne , Versalovic James 
Published: November 2023

BackgroundRecent studies have established that members of the intestinal microbiota communicate with the host central nervous system (CNS) and suggest that this interaction may influence host neural development. Bifidobacterium species in particular are known to produce neuromodulatory metabolites, and previous studies have demonstrated that bifidobacteria can alter CNS gene expression, neurotransmitter function, and rodent behavior. In humans, bifidobacteria are detectable within the first week after birth and are the predominant genus in the infant intestinal microbiota. Given that Bifidobacterium spp. colonize the human system during a critical period of neural circuit development and organization, the Bifidobacterium‐gut‐brain interaction may have pervasive and lasting effects on brain function and behavior. However, we currently lack detailed knowledge regarding which CNS genes are altered by the presence of these species, and how modulation of these genes may affect neurodevelopment. We hypothesized that colonization with a representative Bifidobacterium species would alter CNS expression of genes important to synaptic plasticity and neurodevelopment.Method & ResultsGerm‐free mice were utilized to examine the effects of Bifidobacterium colonization. Adult male and female germ‐free mice were colonized with a representative species (B. dentium) by oral gavage. Age‐matched Specific Pathogen Free (SPF) mice that had a complex murine gut microbiome were also tested for comparison. We found that B. dentium colonization resulted in alterations to intestinal morphology, increased mucus production, and increased luminal concentrations of serotonin in the ileum. Due to its simple, well‐defined, and conserved circuit, the cerebellum was used as a model to study how the microbiota alters CNS gene expression and circuit development. Cerebellar gene expression was quantified via qRT‐PCR. Significant expression differences were observed in 10 of 30 synaptic plasticity‐related genes tested. Mono‐association with B. dentium resulted in a gene expression profile that resembled that of SPF mice for many of the genes tested, suggesting a rescue effect. Based on the qRT‐PCR results, 5 genes of interest were selected to examine via RNA In Situ Hybridization to further quantify and localize expression differences in whole brain sections. These results support the qRT‐PCR data, and have identified other brain regions of interest that exhibit altered gene expression. The marble bury test was utilized to assess variations in stereotypic digging behavior and indicated that B. dentium‐colonized mice displayed altered behavior relative to their SPF and germ‐free counterparts.ConclusionTogether these data demonstrate that mono‐association of germ‐free mice with B. dentium alters intestinal morphology and neurochemical concentrations, as well as CNS gene expression which may correspond with the observed behavioral changes.
MiR9 is increased in Rheumatoid Arthritis Monocytes and Regulates Monocyte Migration
Authors: Gaudette Joshua , Stinson W Alexander , Fox David A , Amin M Asif , Rabquer Bradley James 
Published: November 2023

PurposeRheumatoid Arthritis (RA) is characterized by inflammation and joint degradation. Monocytes, a key leukocyte in inflammation, migrate from the blood to damaged tissues where they differentiate into macrophages that serve multiple immune functions. MicroRNA (miRNA), small noncoding regions of RNA, act as regulators of gene expression, bind to and degrade mRNA. We have previously shown that miR9 is up regulated in THP1 monocytes and U937 monocytes with TNFα stimulation. Here we hypothesize that miR9 is up regulated in the inflammatory disease RA, and that mir9 regulates monocyte migration.Methods and ResultsMonocytes were isolated from blood of patients with RA (N=3) and normal subjects (N=6). MiRNA was isolated, cDNA was prepared, and qPCR was performed using primers specific to miR9. miR9 was significantly upregulated in RA monocytes by 2.3 fold compared to normal monocytes (P < 0.05). We searched for miR9 gene targets using miRWALK, MiRanda, and NCBI databases and found 8,778 gene targets. Targets were narrowed to biological pathways specific to human monocytes and inflammation, leaving 39 gene targets. Of these 39 targets, suppressor of cytokine signaling 4 (SOCS 4) was chosen for further study because it showed the highest seed length for miRNA binding. An inhibitor and mimic for miR9 were used to assess its ability in modulating SOCS4 expression. Results showed a 0.2 fold reduction in expression with the miR9 mimic, and a negligible change in expression from the miR9 inhibitor. Chemotaxis assays were performed on THP‐1 monocytes transfected with miR9 mimic or inhibitor to measure the effect of miR9 on monocyte migration. THP‐1 cells treated with the miR9 mimic displayed an increase in migration by 40% (n=10 replicates) compared to control cells, whereas the inhibitor showed no change in migration (n=10 replicates).ConclusionOur results suggest miR9 is upregulated in RA monocytes and that miR9 plays a role in monocyte migration by regulating genes such as SOCS4.Support or Funding InformationHewlett‐Mellon Fund for Faculty Development at Albion College
Survey on microRNAs Targeting MuRF1 and MuRF2 in Regenerating Skeletal Muscle
Authors: Silva William Jose , Baptista Igor Luchini , Silvestre João Guilherme , Yan Chao Irene , Moriscot Anselmo Sigari 
Published: November 2023

IntroductionSkeletal muscle is a very adaptable tissue, mainly due to its regenerative capacity, which is fundamental importance for repairing large lesions and microlesions as well. Recently our research group observed that the E3 ligases MuRF1 and MuRF2, in addition to being implicated the atrophy process is also essential for muscle regeneration (data not published). In view of these results, in this study we investigated the molecular mechanisms involved in the increased expression of MuRFs during regeneration. Our hypothesis contemplates that certain microRNAs (miRNAs or miRs, regulators of gene expression at post‐transcriptional level) can regulate MuRFs expression during the regeneration process.ObjectiveSearch miRNAs that targets MuRFs, and investigate its impact on the skeletal muscle regeneration.Materials and MethodsC57/BL6 mice (~8wk. 30±3g) were injected with cardiotoxin (CTX, 10μM) in the Tibialis Anterior (TA) muscle for 1, 3 and 10 days in order to induce muscle injury. After these periods the animals were euthanized and the TA muscles were removed for determination of RNA and protein expression levels. Plasmids pMIR‐3UTRMuRF1 and pMIR‐3UTRMuRF2 were produced out of the amplification of the 3′ UTR of each cDNA and cloned into the vector pMIR‐Report system. Vectors (300ng) were co‐transfected with the specified miRNA (miRNA mimics 40uM or mimic Control#1) in HeLa cells. Then, the luciferase assay was performed. Primary myoblast cultures were transfected with siRNA to MuRFs in pre differentiation and post differentiation phases. Procedures and experimental protocols were approved by the Ethics experimental Committee at University of Sao Paulo ‐ CEUA #118.ResultsThe protein expression of MuRF1 rises after 3 days of injury (~100%; p<0.05 vs control), while its gene expression rises as soon as 1 day (~150%; p<0.05 vs control) after injury. MuRF2, gene expression and protein content, rises after 10 days of lesion (~ 50%; p <0.05 vs. control). Transcription factors of MuRFs, FOXO3a and NF‐Kb increase their protein expression after 3 days of injury (>100%; p<0.05 vs. control). The miRs 29c and 101a which target MuRF1 and miRs 133a and 133b which target MuRF2 (TargetScan6.2 analysis) are reduced by 1 and 3 days(~50–80%; p<0.05 vs. control). By using luciferase assays with co‐transfection pMIR‐3UTR‐MuRF1 or pMIR‐3UTR‐MuRF2 and their respective mimics, we showed that miR29c targets MuRF1 and miR133a and 133b target MuRF2. Interestingly, MuRF1 and MuRF2 siRNA knock down do not change the expected modulation of miRs 29c, 133a and 133b. Expression of miRs 133a and 133b are increased after the post differentiation period (~100%; p<0.05 vs pre‐differentiation) in cultured myoblasts.ConclusionWith these partial results, we identify and validate the miRs 29c and 133a/b target MuRFs.Support or Funding InformationGrant 2012/22488‐2, Sao Paulo Research Foundation (FAPESP)
Mechanics of cell motility within stratified epithelia
Authors: Czirok Andras , Isai Dona G , Neufeld Zoltan 
Published: November 2023

Recently we proposed a three dimensional mechanical model of embryonic tissue dynamics. Mechanically coupled adherent cells are represented as particles interconnected with elastic beams which can exert non‐central forces and torques. Tissue plasticity is modeled by a stochastic process consisting of a connectivity change (addition or removal of a single link) followed by a complete relaxation to mechanical equilibrium. In particular, we assume that (i) two non‐connected, but adjacent particles can form a new link; and (ii) the lifetime of links is reduced by tensile forces. This model yields a realistic macroscopic elasto‐plastic behavior and microscopic parameter values can be inferred from tissue thickness, macroscopic elastic modulus and the magnitude and dynamics of intercellular adhesion forces. In addition to their mechanical role, model particles can also act as simulation agents and actively modulate their connectivity according to specific rules. We demonstrate, that anisotropic link insertion and removal probabilities can give rise to local cell intercalation and large scale convergent extension movements. Cell extrusion from an epithelial layer involves mechanical torques, and can generate bending moments within a stratified (multi‐layered) epithelium ‐‐ a process likely to shape the early embryonic heart field.Support or Funding InformationR01‐GM102801
Musculoskeletal and Neural Comparison of Normal and Abnormal Development in Humans and Broader Medical Implications
Authors: Alghamdi Malak , Smith Christopher , Ziermann Janine , Diogo Rui 
Published: November 2023

Anatomical and medical studies of gross anatomical birth defects tend to focus mainly on external features, or in hard tissues (i.e., bones, cartilages, teeth). As a new, long term project we have been collecting and systematically comparing detailed musculoskeletal and neural information on normal human development and phenotypes seen in human fetuses, neonates and adults with conditions as diverse as anencephaly, trisomy 13, trisomy 18, trisomy 21, and/or cyclopia. This comparison is based on careful dissections and an extensive review of the scarce literature on the subject. Our dissections and comparisons revealed that certain similar anatomical patterns are frequently found in each of these conditions, supporting the “logic of monsters” hypothesis of Pere Alberch, according to which even in extreme cases of birth defects (e.g. anencephaly, cyclopia) there is still a discernable, marked “order”, instead of random, chaotic phenotypes, because of strong internal constraints limiting the amount of possible developmental outcomes. For instance, the absence of muscles that are normally polymorphic in humans (e.g., fibularis tertius, palmaris longus), the absence of tendons to the 5th digit of the hands/feet, and/or the presence of a few ‘atavistic’ muscles, are seen over and over in different cases of abnormal developmental studied by us. We will discuss the broader implications of our results and of the observed patterns by discussing crucial developmental and medical topics such as the links between developmental delay/arrest, ‘atavisms’, variations, anomalies, polymorphisms and the “logic of monsters”.Support or Funding InformationGrants: This work is funded by start‐up funds to RD from Howard University College of Medicine, and by PhD funds from the Saudi Arabia government to MA.
Human Hepatic Progenitor Cells in Hepatic Fibrosis of Elderly Cadavers: Cytokeratin 7 Marks Intermediate Hepatocyte‐Like Cells
Authors: Mak Ki M , Png Chien Yi M 
Published: November 2023

Human hepatic progenitor cells (HPCs) are small oval‐shaped cytokeratin (CK) 7 and CK19 expressing cells that normally reside in a quiescent state in the canals of Hering. In response to liver damage, as in alcoholic liver disease, nonalcoholic fatty liver disease, viral hepatitis, and chronic cholestasis, the HPCs become activated. Being bipotential cells, HPCs can differentiate into either cholangiocytes or hepatocytes; the latter occurs via intermediate hepatocyte‐like cells (IHLCs). IHLCs are polygonal in shape with a size intermediate between HPCs and hepatocytes and express phenotypic markers for biliary lineage and hepatocytic lineage. It has been reported in baboons chronically fed alcohol that an increased number of CK7 immunoreactive IHLCs correlated with the progression to more severe stages of liver fibrosis; CK19 immunoreactive IHLCs, however, were not observed. Since elderly cadavers with diverse causes of death demonstrated progressive stages of liver fibrosis (Mak et al., 2012, Anat Rec 295:40–50), the present study evaluated whether CK7 served as a marker for IHLCs of elderly cadavers. To that end, consecutive paraffin liver sections from embalmed cadavers were immunoperoxidase stained using monoclonal CK7 and CK19 antibodies, in conjunction with EDTA‐based antigen unmasking. For assessment of fibrosis, sections were post‐stained with Sirius red stain for collagens. HPCs were detected by the CK7 or CK19 staining of the cytoplasm, whereas IHLCs were identified by a distinct CK7 staining of the cell membranes and a faint staining of the cytoplasm. In livers with minimal fibrosis, while IHLCs were not observed, CK7 and CK19 positive HPCs were present in both the periportal and lobular parenchyma. However, in livers with perisinusoidal fibrosis, IHLCs were occasionally detectable in foci showing parenchymal fibrosis. In septal fibrosis, IHLCs were readily found, along with CK7 positive HPCs, in close proximity to the developing septa and linking septa. In cirrhosis, there were variable numbers of CK7 stained IHLCs in the parenchyma of the nodules. In striking contrast, CK19 immunoreactive IHLCs were rare or undetectable in any of the abovementioned locations.Conclusionsas in alcoholic fibrosis of baboons, CK7, but not CK19, serves as a marker for IHLCs in liver fibrosis of elderly cadavers. Since IHLCs have the potential to differentiate into hepatocytes, their development in liver fibrosis points to a link between IHLCs and hepatic regeneration consequent to chronic liver injury in the aged cadaveric liver.
Innervation of the Female Levator Ani Muscle and Consequences in Pelvic Floor Dysfunctions
Authors: Bessede Thomas , Timoh Krystel Nyangoh , Lebacle Cedric , Martinovic Jelena , Zaitouna Mazen , Diallo Djibril , Creze Maud , Benoit Gerard , Moszkowicz David 
Published: November 2023

IntroductionThe Levator Ani Muscle (LAM) and the Levator Ani Nerve (LAN) are distended and traumatized during pregnancy and vaginal delivery. Both mechanisms may sustainably impair the pelvis statics but the topography and functions of the nerve supply to the LAM remain anatomically controversial.ObjectiveTo characterize the nature and function of the LAM nerve pathways and to perform a comprehensive 3D reconstruction of the female pelvic innervation.MethodsThe Computer‐Assisted Anatomical dissection (CAAD) protocol was applied to pelvic blocks procured in 6 female human fetuses, in accordance with the National Biomedicine Agency. Specimens were serially sectioned and immunostained for: overall (S100), somatic (PMP22), adrenergic (TH), cholinergic (VaCHT) and nitrergic (nNOS) nerve fibers and for: smooth (smooth actin) and striated (myogenin) muscle fibers. Slides were digitized for 3D reconstructions on WinSurf®. A pelvic sample of an additional fetus was examined under electronic microscopy to analyze the ultrastructure of the LAM.ResultsThree main nerve pathways to the LAM were observed: the levator ani nerve (LAN), the pudendal nerve (PuN), the inferior hypogastric plexus (IHP). The PuN was both somatic and autonomic, located below the LAM (‘infralevator pathway’) and supplying innervation to the inferior aspect of the LAM. The LAN was solely somatic, located above the LAM (‘supralevator pathway’) and supplying innervation to the superior aspect of the LAM. The IHP nerve fibers were solely autonomic, located in between the LAM and the pelvic organs (‘endolevator pathway’) and supplying innervation to the medial portion of the LAM. Smooth muscle areas were identified with labelings and electronic microscopy: they 1) were located in the puboanal sling of the LAM 2) were bulging between rectum and vagina 3) were intermingled to the anococcygeal raphe 4) were innervated by the IHP and the PuN pathways.ConclusionHaving at once striated and smooth muscle fibers and being co‐innervated by the somatic and autonomic systems, the LAM represents a crucial anatomical structure involved in the pelvic floor function and its rehabilitation.Support or Funding Informationnone
Transcriptome profiles of porcine skin dyschromia after wound injury
Authors: Alkhalil Abdulnaser , Muhie Seid , Carney Bonnie , Travis Taryn E , Moffatt Lauren , Johnson Laura , Ramella‐Roman Jessica , Shupp Jeffrey 
Published: November 2023

Dyspigmentation after cutaneous injury negatively impacts patients’ self esteem and social reintegration. Most patients with deep surgical wounds or full thickness burns develop dyspigmented scar tissue which is more salient among people with dark colored skin. Although the pigment synthesis cascade in healthy tissue has been characterized, our understanding of the causative biological perturbations leading to skin dyspigmentation after injury is limited. Consequently, therapies are partially effective, and often invasive and inconvenient. We have adapted a genome‐wide approach to profile the difference in the transcriptomes of hyper‐ and hypo‐pigmented skin obtained from a porcine scar animal model after full thickness wound injury. The results showed significant differences in several genes of chromia including those encoding the DOPA chrome tautomerase (DCT), tyrosinase (TYR), tyrosinase related protein 1 (TYRP‐1), endothelin receptor type B (ENDRB), melanocortin 1 receptor (MC1R) and microphthalmia transcription factor (MITF). Other genes related to melanosome biogenesis such as members of the solute carrier family SLC24A5 and SLC52 A4 and related isotypes including SLC25A19, SLC25A36, SLC35D1, SLC35F5, SLC37A1, SLC52A2 were differentially transcribed. Molecules affecting cell‐matrix and cell‐cell binding showed significant modulations. The Ingenuity pathway analysis revealed the significant alterations to the melanin synthesis pathway and other pathways that indirectly influence melanin synthesis included the Ras signaling and the Wnt/B catanine pathways. The enrichment of several modified biological processes to melanocytes highlights the susceptibility of these cells to the negative impact of deep wounds and worse outcome.Support or Funding InformationThis work was funded by the NIH grant No. 1R15EB013439
Sifting through the Haystack to Find the Needle: Spatial Identification of a Structural Conduit for Transfer of Thermal Activation Energy in the Course of C‐H Cleavage in Soybean Lipoxygenase
Authors: Offenbacher Adam , Hu Shenshen , Nguy Andy , Iavarone Anthony , Klinman Judith 
Published: November 2023

Protein motions are increasingly recognized as an important facet for the huge rate accelerations achieved by enzymes. The ability to link specific protein fluctuations to the chemical step(s) of catalysis remains a daunting challenge. The importance of room temperature hydrogen tunneling in C‐H activation, where the hydrogenic wave function overlap itself is temperature independent, makes such reactions ideal for identifying the thermally activated protein motions that comprise the reaction barrier. We present a thorough examination, involving time, temperature and mutation dependence of the hydrogen deuterium exchange (HDX) behavior of a large, single polypeptide protein, soybean lipoxygenase (SLO‐1), which has emerged as the paradigm tunneling enzyme. By incorporating mutation‐induced packing defects previously shown to selectively impair deuterium over protium tunneling, we first confirm previously proposed changes in active site flexibility as the origin of increases in the temperature dependence of kinetic isotope effects. This approach also identifies a remote surface loop, located approximately 15–40 Å from the active site, whose motions respond to the temperature of the solvent in a manner similar to catalysis. A network that connects this loop to the reactive iron center is identified and concluded to act as a conduit for the funneling of kinetic energy from solvent to the enzyme active site. Our ongoing work is taking advantage of thiol‐based labeling approaches to selectively position chromophores onto the surface loop, to monitor the protein motions and their interactions with solvent molecules in real time from picosecond to microsecond timescales.Support or Funding InformationSupported by a grant from the National Institutes of Health, GM025765 (J.K.) and 1F32GM113432 (A.O.)
ETV5, an Ets Family Transcription Factor, is a Marker for RAS‐Dependent Papillary Thyroid Cancer
Authors: Hoang Nguyet Minh , Puli Oorvashi Roy , Danysh Brian , Hofmann Marie Claude 
Published: November 2023

IntroductionBRAF V600E mutation has a significant association with recurrence in patients with papillary thyroid cancer, reducing survival rate from 98% to 40%. Patients treated with BRAF V600E inhibitors such as vemurafenib acquire resistance to the drug over time (6 months) and tumor cells metastasize to distant organs, leading to patient death. Within the group of patients with BRAFV600E tumors, there are no reliable biomarkers to predict local or distant recurrences, especially when tumors acquire clonal resistance due to the effects of targeted therapies. PTC (Papillary Thyroid Cancer) patients with the BRAF V600E and RAS mutations show an elevated ETV5 (Ets‐transcript variant 5) expression. ETV5, a transcription factor, is known to promote epithelial to mesenchymal transition (EMT) in endometrial carcinomas and ovarian cancer by up‐regulating ZEB1, a repressor of E‐Cadherin, causing cell detachment and invasion. ETV5 also induces the expression of matrix metalloproteinase 2 (MMP‐2) in human endometrial carcinomas, causing invasion.HypothesisWe hypothesize that ETV5 drives cell proliferation and EMT (Epithelial to Mesenchymal Transition) in advanced papillary thyroid cancer.MethodsWe used the cell line KTC1 (BRAF V600E mutant thyroid cancer cell line) as a model. ETV5 and BRAF V600E were knocked‐down using Silencer Select siRNA. Alternatively, cells were treated with standard pharmacological inhibitors (i.e. PI3Ki, AKTi, vemurafenib, ERKi, TGFBR1i). ETV5 expression was quantified using Quantitative PCR and Western Blots. Proliferation/growth assays were performed for 5 days and the cell numbers were counted using the IN Cell 6000 confocal system and Image Analysis software post‐transfection/treatment.ResultsKnockdown of ETV5 and/or BRAF decreased KTC1 cell proliferation. Cell growth was reduced upon treatment with vemurafenib, or PI3K‐AKT and ERK inhibitors, but not with TGFBR1 inhibitors. In comparison to DMSO treatment (control), ETV5 levels were down‐regulated upon ERK inhibitor treatment but were increased when treated with PI3K inhibitors. Short‐term vemurafenib treatment (48h) showed a reduction in ETV5, N‐Cadherin, and CXCR4 expression suggesting its role in EMT.ConclusionETV5 acts downstream of the RAS‐MAPK (ERK1/2) pathway. ETV5 is crucial for KTC1 cell proliferation and may also play a role in EMT in PTC.Support or Funding InformationThis project is sponsored by a DRP grant, which is part of NIH P50 CA168505