EFFECT OF THE C-TERMINUS OF TSHR ON T4 AND TRAB EXPRESSION IN BALB/C MICE.

Context: The expression of TSHR-C on the serum tetraiodothyronine (T4) and TSH receptor antibody (TRAb) levels are rarely studied. Objective: The effect of TSHR-c on T4 and TRAb levels and concomitant thyroid histological changes in mice was investigated. Design: Animal experimental study. Subjects and methods: Female BALB/c mice at 6-8 weeks of age were immunized with the thyroid stimulating hormone receptor antigen C-terminus (TSHR-C), and randomly divided into control group (treated with the corresponding concentrations of normal saline) and four experimental subgroups: TSHR-c1 subgroup (4 µg), TSHR-c2 subgroup (6 µg), TSHR-c3 subgroup (8 µg) and TSHR-c4 subgroup (10 µg). Serum T4 and TRAb levels were determined. Results: The serum T4 level decreased significantly in the experimental mice as the concentration increased. All the experimental mice were positive for serum TRAb (experimental groups: 40 positive/40, 100% vs. control group: 3 positive/10, 30%) compared to the control group (P =0.000). HE staining showed that the follicles in the control mice were composed of small to medium-sized round follicles, whereas the follicles in the experimental mice were irregularly enlarged under light microscope. Conclusions: TSHR-c immunization resulted in thyroid hormone changes like those observed in hypothyroidism, probably due to the induction of TRAb generation.

Patient motivation and satisfaction from orthognathic surgery-a case series.

OBJECTIVE: This study attempted to understand the specific motivations of patients who undergo orthognathic surgery and determine their satisfaction with the surgery. The study also assessed the prevalence of complications and how they may impact patient satisfaction. STUDY DESIGN: Patients who underwent orthognathic surgery at the University of California between 2016 and 2019 and had completed postoperative orthodontic treatment for ≥9 months were interviewed. They responded to an open-ended telephone interview regarding their motivations, satisfaction, and complications. RESULTS: The patients showed a high level of satisfaction with the surgery, but there were persistent complications that affected satisfaction. The predominant complication was paresthesia over the distribution of the inferior alveolar nerve. The majority of patients who reported prior headaches and temporomandibular joint problems described improvement in those areas. Comparing the patients' motivations before and after surgery showed that before surgery, patients reported functional concerns, whereas postoperatively they were much more likely to recall aesthetic reasons for the surgery. CONCLUSION: This study showed that although patients are generally satisfied after orthognathic surgery, patients need to be realistically informed of their expectations and adequately informed of possible complications.

Mutation-specific pathophysiological mechanisms define different neurodevelopmental disorders associated with SATB1 dysfunction.

Whereas large-scale statistical analyses can robustly identify disease-gene relationships, they do not accurately capture genotype-phenotype correlations or disease mechanisms. We use multiple lines of independent evidence to show that different variant types in a single gene, SATB1, cause clinically overlapping but distinct neurodevelopmental disorders. Clinical evaluation of 42 individuals carrying SATB1 variants identified overt genotype-phenotype relationships, associated with different pathophysiological mechanisms, established by functional assays. Missense variants in the CUT1 and CUT2 DNA-binding domains result in stronger chromatin binding, increased transcriptional repression, and a severe phenotype. In contrast, variants predicted to result in haploinsufficiency are associated with a milder clinical presentation. A similarly mild phenotype is observed for individuals with premature protein truncating variants that escape nonsense-mediated decay, which are transcriptionally active but mislocalized in the cell. Our results suggest that in-depth mutation-specific genotype-phenotype studies are essential to capture full disease complexity and to explain phenotypic variability.

Advances in the development of gene therapy, noncoding RNA, and exosome-based treatments for tendinopathy.

Tendinopathy is a common musculoskeletal disorder characterized by chronic low-grade inflammation and tissue degeneration. Tendons have poor innate healing ability and there is currently no cure for tendinopathy. Studies elucidating mechanisms underlying the pathogenesis of tendinopathy and mechanisms mediating the genesis of tendons during development have provided novel targets and strategies to enhance tendon healing and repair. This review summarizes the current understanding and treatments for tendinopathy. The review also highlights recent advances in gene therapy, the potential of noncoding RNAs, such as microRNAs, and exosomes, which are nanometer-sized extracellular vesicles secreted from cells, for the treatment of tendinopathy.

De novo variants in CELF2 that disrupt the nuclear localization signal cause developmental and epileptic encephalopathy.

We report heterozygous CELF2 (NM_006561.3) variants in five unrelated individuals: Individuals 1-4 exhibited developmental and epileptic encephalopathy (DEE) and Individual 5 had intellectual disability and autistic features. CELF2 encodes a nucleocytoplasmic shuttling RNA-binding protein that has multiple roles in RNA processing and is involved in the embryonic development of the central nervous system and heart. Whole-exome sequencing identified the following CELF2 variants: two missense variants [c.1558C>T:p.(Pro520Ser) in unrelated Individuals 1 and 2, and c.1516C>G:p.(Arg506Gly) in Individual 3], one frameshift variant in Individual 4 that removed the last amino acid of CELF2 c.1562dup:p.(Tyr521Ter), possibly resulting in escape from nonsense-mediated mRNA decay (NMD), and one canonical splice site variant, c.272-1G>C in Individual 5, also probably leading to NMD. The identified variants in Individuals 1, 2, 4, and 5 were de novo, while the variant in Individual 3 was inherited from her mosaic mother. Notably, all identified variants, except for c.272-1G>C, were clustered within 20 amino acid residues of the C-terminus, which might be a nuclear localization signal. We demonstrated the extranuclear mislocalization of mutant CELF2 protein in cells transfected with mutant CELF2 complementary DNA plasmids. Our findings indicate that CELF2 variants that disrupt its nuclear localization are associated with DEE.

A Converging Strategy for the Generation of a Virtually Sequenced cDNA Library from Unreferenced Pacific Oysters.

The access to biological material of reference species, which were used previously in key experiments such as in the development of novel cell lines or genome sequencing projects, are often difficult to provide for further studies or third parties due to the consumptive nature of the samples. Although now widely distributed over the Pacific coasts of Asia, Australia and North America, individual Pacific oyster specimens are genetically quite diverse and are therefore not directly suitable as the starting material for gene libraries. In this article, we demonstrate the use of unreferenced Pacific oyster specimens obtained from regional seafood markets to generate cDNA libraries. These libraries were then compared to the publicly available oyster genome, and the closest related library was selected using the mitochondrial reference genes Cytochrome C Oxidase subunit I (COX1) and NADH Dehydrogenase (ND). The suitability of the generated cDNA library is also demonstrated by cloning and expression of two genes encoding the enzymes UDP-glucuronic acid dehydrogenase (UGD) and UDP-xylose synthase (UXS), which are responsible for the biosynthesis of UDP-xylose from UDP-glucose.

CITED2 mediates the cross-talk between mechanical loading and IL-4 to promote chondroprotection.

Osteoarthritis (OA) pathogenesis is mediated largely through the actions of proteolytic enzymes such as matrix metalloproteinase (MMP) 13. The transcriptional regulator CITED2, which suppresses the expression of MMP13 in chondrocytes, is induced by interleukin (IL)-4 in T cells and macrophages, and by moderate mechanical loading in chondrocytes. We tested the hypothesis that CITED2 mediates cross-talk between IL-4 signaling and mechanical loading-induced pathways that result in chondroprotection, at least in part, by downregulating MMP13. IL-4 induced CITED2 gene expression in human chondrocytes in a dose- and time-dependent manner through JAK/STAT signaling. Mechanical loading combined with IL-4 resulted in additive effects on inducing CITED2 expression and downregulating of MMP13 in human chondrocytes in vitro. In vivo, IL-4 gene knockout (KO) mice exhibited reduced basal levels of CITED2 expression in chondrocytes. While moderate treadmill running induced CITED2 expression and reduced MMP13 expression in wild-type mice, these effects were blunted (for CITED2) or abolished (for MMP13) in chondrocytes of IL-4 gene KO mice. Moreover, intra-articular injections of mouse recombinant IL-4 combined with regular cage activity mitigated post-traumatic OA to a greater degree compared to immobilized mice treated with IL-4 alone. These data suggest that using moderate loading to enhance IL-4 may be a potential therapeutic strategy for chondroprotection in OA.

CITED2 mediates the mechanical loading-induced suppression of adipokines in the infrapatellar fat pad.

Adipokines secreted from the infrapatellar fat pad (IPFP), such as adipsin and adiponectin, have been implicated in osteoarthritis pathogenesis. CITED2, a mechanosensitive transcriptional regulator with chondroprotective activity, may modulate their expression. Cited2 haploinsufficient mice (Cited2+/- ) on a high-fat diet (HFD) exhibited increased body weight and increased IPFP area compared to wild-type (WT) mice on an HFD. While an exercise regimen of moderate treadmill running induced the expression of CITED2, as well as PGC-1α, and reduced the expression of adipsin and adiponectin in the IPFP of WT mice on an HFD, Cited2 haploinsufficiency abolished the loading-induced expression of PGC-1α and loading-induced suppression of adipsin and adiponectin. Furthermore, knocking down or knocking out CITED2 in adipose stem cells (ASCs)/preadipocytes derived from the IPFP in vitro led to the increased expression of adipsin and adiponectin and reduced PGC-1α, and abolished the loading-induced suppression of adipsin and adiponectin and loading-induced expression of PGC-1α. Overexpression of PGC-1α in these ASC/preadipocytes reversed the effects caused by CITED2 deficiency. The current data suggest that CITED2 is a critical regulator in physiologic loading-induced chondroprotection in the context of an HFD and PGC-1α is required for the inhibitory effects of CITED2 on the expression of adipokines such as adipsin and adiponectin in the IPFP.

Epigallocatechin-3-gallate suppresses pain-related and proinflammatory mediators in the subacromial bursa in rotator cuff tendinopathy.

Rotator cuff tendinopathy is one of the leading causes of shoulder pain. However, the mechanisms involved in the development of rotator cuff tendinopathy pain are not fully understood. In this study, we first examined the histological features of subacromial bursa from patients with rotator cuff tendinopathy who had symptoms of pain, and investigated the expression of pain mediators, proinflammatory cytokines, metalloproteinases, growth factors, and alarmins in diseased tendon and bursa tissue by real-time PCR, western blot, and/or immunohistochemistry/immunofluorescence staining. Then we investigated whether epigallocatechin-3-gallate (EGCG) could reduce the expression of pain mediators and proinflammatory cytokines in human primary bursa cells and explored the paracrine effect of these EGCG-treated bursa cells on tenocytes in vitro. Neovascularization and infiltration of immune cells including monocytes/macrophages and mast cells were observed in diseased bursa tissue. Bursa from patients with pain had higher mRNA expression of pain mediators and proinflammatory cytokines, compared to the rotator cuff tendon of the same patients, as well as the bursa from asymptomatic patients. EGCG treatment significantly suppressed the interleukin 1 beta (IL-1ß)-induced elevation of mRNA expression of pain mediators, proinflammatory cytokines, and matrix metalloproteinases (MMPs) in bursa cells in vitro; conditioned medium from EGCG-treated bursa cells significantly reduced IL-1ß-induced expression in human primary tenocytes. Our study suggests that the subacromial bursa might serve as a local source of pain mediators and proinflammatory cytokines in rotator cuff tendinopathy. Moreover, EGCG treatment by primarily targeting the subacromial bursa may be a potential strategy to relieve rotator cuff tendinopathy-related pain and symptoms.

Correction to: Green tea polyphenol treatment is chondroprotective, anti-inflammatory and palliative in a mouse posttraumatic osteoarthritis model.

Following publication of the original article [1], the authors reported an error in Figs. 2C and 5C.

Role of pannexin 1 channels in load-induced skeletal response.

The pannexin 1 (Panx1) channel is a mechanosensitive channel that interacts with P2X7 receptors (P2X7R) to form a functional complex that has been shown in vitro to play an essential role in osteocyte mechanosignaling. While the participation of P2X7R in skeletal responses to mechanical loading has been demonstrated, the role of Panx1 and its interplay with P2X7R still remain to be determined. In this study, we use a global Panx1-/- mouse model and in vivo mechanical loading to demonstrate that Panx1 channels play an essential role in load-induced skeletal responses. We found that absence of Panx1 not only disrupts the P2X7R-Panx1 signaling complex, but also alters load-induced regulation of P2X7R expression. Moreover, lack of Panx1 completely abolished load-induced periosteal bone formation. Load-induced regulation of ß-catenin and sclerostin expression was dysregulated in Panx1-/- , compared to wild-type, bone. This finding suggests that Panx1 deficiency disrupts Wnt/ß-catenin signaling by lowering ß-catenin while favoring inhibition of bone formation by increasing load-induced sclerostin expression. This study demonstrates the existence of a Panx1-dependent mechanosensitive mechanism that not only modulates ATP signaling but also coordinates Wnt/ß-catenin signaling that is essential for proper skeletal response to mechanical loading.

Retraction Note: The Role of Scleraxis in Fate Determination of Mesenchymal Stem Cells for Tenocyte Differentiation.

This paper has been retracted.

Nutraceuticals and osteoarthritis pain.

Arthritis is a chronic disease of joints. It is highly prevalent, particularly in the elderly, and is commonly associated with pain that interferes with quality of life. Because of its chronic nature, pharmacological approaches to pain relief and joint repair must be safe for long term use, a quality many current therapies lack. Nutraceuticals refer to compounds or materials that can function as nutrition and exert a potential therapeutic effect, including the relief of pain, such as pain related to arthritis, of which osteoarthritis (OA) is the most common form. Of interest, nutraceuticals have recently been shown to have potential in relieving OA pain in human clinical trials. Emerging evidence indicates nutraceuticals may represent promising alternatives for the relief of OA pain. In this paper, we will overview OA pain and the use of nutraceuticals in OA pain management, focusing on those that have been evaluated by clinical trials. Furthermore, we discuss the biologic and pharmacologic actions underlying the nutraceutical effects on pain relief based on the potential active ingredients identified from traditional nutraceuticals in OA pain management and their potential for drug development. The review concludes by sharing our viewpoints that future studies should prioritize elucidating the mechanisms of action of nutraceuticals in OA and developing nutraceuticals that not only relieve OA pain, but also mitigate OA pathology.

Exosomes: biology, therapeutic potential, and emerging role in musculoskeletal repair and regeneration.

Exosomes are nanovesicles secreted from cells that play key roles in intercellular communication. They carry unique content derived from parental cells and are capable of transferring this cargo between cells. The role and function of exosomes largely depends on the origin and functional status of the parental cells. Emerging evidence indicates that exosomes are associated with biological processes and pathogenesis of certain diseases. These nanovesicles offer great potential as biomarkers, enabling the monitoring and diagnosis of various diseases in a noninvasive manner. Furthermore, as an efficient vehicle of biomolecular intercellular transfer, exosomes are under intensive investigation for their potential for drug delivery and carriers for gene therapy. Here, we first summarize the basic biology and function of exosomes, followed by a discussion of their clinical potential, including the use of exosomes for disease diagnosis, treatment, and drug delivery. The review will highlight the potential of exosomes derived from stem cells in regenerative medicine, with a focus on musculoskeletal tissues. We conclude by sharing our views on the challenges, opportunities, and future directions for the use of exosomes as a therapeutic treatment for the repair and regeneration of musculoskeletal tissues.

Extended vs. brief intermittent access to palatable food differently promote binge-like intake, rejection of less preferred food, and weight cycling in female rats.

BACKGROUND: Palatable food access promotes obesity leading some to diet. Here, we modeled the roles of duration, intermittency and choice of access in bingeing, escalation of daily intake, and underacceptance of alternatives. METHOD: Female rats with ("Choice") or without continuous chow access, received chow or continuous (Chocolate), intermittent (MWF) long (24h, Int-Long), or intermittent short (30min, Int-Short) access to a sucrose-rich, chocolate-flavored diet (CHOC). RESULTS: Int-Long rats showed cycling body weight; they overate CHOC, had increased feed efficiency on access days and underate chow and lost weight on non-access days, the latter correlating with their reduced brown fat. Int-Short rats had the greatest 30-min intake upon CHOC access, but did not underaccept chow or weight cycle. Individual vulnerability for intermittent access-induced feeding adaptations was seen. Continuous access rats gained fat disproportionate, but in direct relation, to their normalized energy intake and persistently underaccepted chow despite abstinence and return to normal weight. Abstinence reduced the binge-like CHOC intake of Int-Short rats and increased that of continuous access rats, but not to levels associated with intermittent access history. Choice increased daily CHOC intake under Continuous access and binge-like intake under Int-Short access. CONCLUSIONS: Intermittency and duration of past access to palatable food have dissociable, individually-vulnerable influences on its intake and that of alternatives. With extended access, daily intake reflects the palatability of available food, rather than metabolic need. Ongoing restrictedness of access or a history of intermittency each drive binge-like intake. Aspects of palatable food availability, similar and different to drug availability, promote disordered eating.

Advances of stem cell based-therapeutic approaches for tendon repair.

Tendon injuries are significant clinical problems. Current treatments often result in incomplete repair or healing, which may lead to reduced function and rupture. Stem cell-based therapy is a promising intervention for tendon repair. In this article, we attempt to provide a brief overview on the recent progress in the field, current understanding of the underlying mechanisms of the approach, and the potential of stem cell-based therapies beyond cell implantation. We conclude the review by sharing our viewpoints on the challenges, opportunities, and future directions of this approach. The translational potential of this article: This paper reviews recent progress on stem cell-based therapeutic approaches for tendon repair, which highlights its translational potential and challenges.

Procyanidins Mitigate Osteoarthritis Pathogenesis by, at Least in Part, Suppressing Vascular Endothelial Growth Factor Signaling.

Procyanidins are a family of plant metabolites that have been suggested to mitigate osteoarthritis pathogenesis in mice. However, the underlying mechanism is largely unknown. This study aimed to determine whether procyanidins mitigate traumatic injury-induced osteoarthritis (OA) disease progression, and whether procyanidins exert a chondroprotective effect by, at least in part, suppressing vascular endothelial growth factor signaling. Procyanidins (extracts from pine bark), orally administered to mice subjected to surgery for destabilization of the medial meniscus, significantly slowed OA disease progression. Real-time polymerase chain reaction revealed that procyanidin treatment reduced expression of vascular endothelial growth factor and effectors in OA pathogenesis that are regulated by vascular endothelial growth factor. Procyanidin-suppressed vascular endothelial growth factor expression was correlated with reduced phosphorylation of vascular endothelial growth factor receptor 2 in human OA primary chondrocytes. Moreover, components of procyanidins, procyanidin B2 and procyanidin B3 exerted effects similar to those of total procyanidins in mitigating the OA-related gene expression profile in the primary culture of human OA chondrocytes in the presence of vascular endothelial growth factor. Together, these findings suggest procyanidins mitigate OA pathogenesis, which is mediated, at least in part, by suppressing vascular endothelial growth factor signaling.

Mesenchymal stem cells in tendon repair and regeneration: basic understanding and translational challenges.

Tendon injuries are common and present a clinical challenge because they often respond poorly to treatment and require prolonged rehabilitation. Current treatments often do not completely repair or regenerate the injured or diseased tendon to its native composition, structure, and mechanical properties. Stem cell-based therapies have brought new hope for tissue repair and regeneration, including that for tendon rupture and tendinopathy. Despite tremendous effort and progress, the success of stem cell-based studies on tendon repair and regeneration has mainly been limited to preclinical studies with few clinical applications. In this concise review, we discuss basic understanding and translational challenges of using mesenchymal stem cells (MSCs) for tendon repair and regeneration, with a focus on (1) tendon stem/progenitor cells (TSPCs) and therapeutic approaches using TSPCs and other MSCs, (2) regulation of fate determination in MSCs for tendon-lineage differentiation, (3) pretreatment and condition of stem/progenitor cells for transplantation, and (4) a treatment approach that involves stimulating endogenous stem cells to enhance tendon healing. The review concludes with discussion on future directions.

Curcumin slows osteoarthritis progression and relieves osteoarthritis-associated pain symptoms in a post-traumatic osteoarthritis mouse model.

BACKGROUND: Curcumin has been shown to have chondroprotective potential in vitro. However, its effect on disease and symptom modification in osteoarthritis (OA) is largely unknown. This study aimed to determine whether curcumin could slow progression of OA and relieve OA-related pain in a mouse model of destabilization of the medial meniscus (DMM). METHODS: Expression of selected cartilage degradative-associated genes was evaluated in human primary chondrocytes treated with curcumin and curcumin nanoparticles and assayed by real-time PCR. The mice subjected to DMM surgery were orally administered curcumin or topically administered curcumin nanoparticles for 8 weeks. Cartilage integrity was evaluated by Safranin O staining and Osteoarthritis Research Society International (OARSI) score, and by immunohistochemical staining of cleaved aggrecan and type II collagen, and levels of matrix metalloproteinase (MMP)-13 and ADAMTS5. Synovitis and subchondral bone thickness were scored based on histologic images. OA-associated pain and symptoms were evaluated by von Frey assay, and locomotor behavior including distance traveled and rearing. RESULTS: Both curcumin and nanoparticles encapsulating curcumin suppressed mRNA expression of pro-inflammatory mediators IL-1ß and TNF-α, MMPs 1, 3, and 13, and aggrecanase ADAMTS5, and upregulated the chondroprotective transcriptional regulator CITED2, in primary cultured chondrocytes in the absence or presence of IL-1ß. Oral administration of curcumin significantly reduced OA disease progression, but showed no significant effect on OA pain relief. Curcumin was detected in the infrapatellar fat pad (IPFP) following topical administration of curcumin nanoparticles on the skin of the injured mouse knee. Compared to vehicle-treated controls, topical treatment led to: (1) reduced proteoglycan loss and cartilage erosion and lower OARSI scores, (2) reduced synovitis and subchondral plate thickness, (3) reduced immunochemical staining of type II collagen and aggrecan cleavage epitopes and numbers of chondrocytes positive for MMP-13 and ADAMTS5 in the articular cartilage, and (4) reduced expression of adipokines and pro-inflammatory mediators in the IPFP. In contrast to oral curcumin, topical application of curcumin nanoparticles relieved OA-related pain as indicated by reduced tactile hypersensitivity and improved locomotor behavior. CONCLUSION: This study provides the first evidence that curcumin significantly slows OA disease progression and exerts a palliative effect in an OA mouse model.

Rho/Rock signal transduction pathway is required for MSC tenogenic differentiation.

Mesenchymal stem cell (MSC)-based treatments have shown promise for improving tendon healing and repair. MSCs have the potential to differentiate into multiple lineages in response to select chemical and physical stimuli, including into tenocytes. Cell elongation and cytoskeletal tension have been shown to be instrumental to the process of MSC differentiation. Previous studies have shown that inhibition of stress fiber formation leads MSCs to default toward an adipogenic lineage, which suggests that stress fibers are required for MSCs to sense the environmental factors that can induce differentiation into tenocytes. As the Rho/ROCK signal transduction pathway plays a critical role in both stress fiber formation and in cell sensation, we examined whether the activation of this pathway was required when inducing MSC tendon differentiation using rope-like silk scaffolds. To accomplish this, we employed a loss-of-function approach by knocking out ROCK, actin and myosin (two other components of the pathway) using the specific inhibitors Y-27632, Latrunculin A and blebbistatin, respectively. We demonstrated that independently disrupting the cytoskeleton and the Rho/ROCK pathway abolished the expression of tendon differentiation markers and led to a loss of spindle morphology. Together, these studies suggest that the tension that is generated by MSC elongation is essential for MSC teno-differentiation and that the Rho/ROCK pathway is a critical mediator of tendon differentiation on rope-like silk scaffolds.