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Item Efficacy and mechanism of Wuzi Yanzong pill on the prevention and treatment of EAE(Cell Press, 2023-10-04) Li, Yan-Rong; Zhang, Ruo-Nan; Sun, Rui-Rui; Li, Yan-Yan; Zhang, Bo; Jin, Xiao-Ming; Zhang, Hai-Fei; Xiao, Bao-Guo; Ma, Cun-Gen; Fan, Hui-Jie; Chai, Zhi; Anatomy, Cell Biology and Physiology, School of MedicineObjective: Studies have shown that Wuzi Yanzong Pill (WYP) can be used to treat neurological diseases, but its mechanisms for multiple sclerosis (MS) remain unclear. This study aims to determine the effect of WYP on MS in an animal model of experimental autoimmune encephalomyelitis (EAE), and explore its mechanism. To provide theoretical basis for the clinical treatment of MS with WYP. Methods: C57BL/6 female mice were randomly divided into Blank control, EAE control, low dose WYP, medium dose WYP, and high dose WYP groups. One week before model generation, the mice were gavaged with saline (50 mL/kg/d) in Blank control and EAE control groups. The treatment groups was gavaged with different doses of WYP solution (4, 8, or 16 g/kg/d respectively) Clinical scores were recorded daily. Sample collection was conducted on the 14th and 28th days, respectively The expressions of IL-10, IL-17, IL-12, TNF-α and IFN-γ in spleen were detected by ELISA. The expressions of ROCKII, P-MYPT1, TLR4, NF-κB/p65, MCP-1, CCR2 in spleen, brain and spinal cord were detected by Western Blot. The types of macrophages and the contents of intracellular IL-10 and IL-12 were detected by Flow Cytometry. The contents of TNF-α and TLR4 mRNA in the spleen were detected by RT-PCR. Results: WYP treatment improved the clinical score of EAE mice in a significant dose-dependent manner, with the WYP high-dose group showed the most significant improvement in clinical score. Compared with the EAE control group, WYP high dose group had significantly lower levels of IL-17, IFN-γ, ROCKII, P-MYPT1, TLR4, NF-κB/p65, MCP-1, and CCR2 as well as TNF-α and TLR4 mRNA, but increased the number of M2 macrophages and IL-10. Conclusion: WYP treatment relieves clinical symptoms in EAE mice, which may be related to regulate inflammatory pathway and inhibiting expressions of inflammatory cytokines.Item Unilateral Hypofunction of the Masseter Leads to Molecular and 3D Morphometric Signs of Atrophy in Ipsilateral Agonist Masticatory Muscles in Adult Mice(MDPI, 2023-09-29) Balanta-Melo, Julián; Eyquem-Reyes, Andrea; Blanco, Noelia; Vásquez, Walter; Kupczik, Kornelius; Toro-Ibacache, Viviana; Buvinic, Sonja; Anatomy, Cell Biology and Physiology, School of MedicineMice are commonly used to study mandibular dynamics due to their similarity in chewing cycle patterns with humans. Adult mice treated unilaterally with botulinum toxin type A (BoNTA) in the masseter exhibit atrophy of this muscle characterized by an increase in the gene expression of atrophy-related molecular markers, and a reduction in both muscle fiber diameter and muscle mass at 14d. However, the impact of this muscle imbalance on the non-treated masticatory muscles remains unexplored. Here, we hypothesize that the unilateral masseter hypofunction leads to molecular and 3D morphometric signs of atrophy of the masseter and its agonist masticatory muscles in adult mice. Twenty-three 8-week-old male BALB/c mice received a single injection of BoNTA in the right masseter, whereas the left masseter received the same volume of saline solution (control side). Animals were euthanized at 2d, 7d, and 14d, and the masticatory muscles were analyzed for mRNA expression. Five heads were harvested at 14d, fixed, stained with a contrast-enhanced agent, and scanned using X-ray microtomography. The three-dimensional morphometric parameters (the volume and thickness) from muscles in situ were obtained. Atrogin-1/MAFbx, MuRF-1, and Myogenin mRNA gene expression were significantly increased at 2 and 7d for both the masseter and temporalis from the BoNTA side. For medial pterygoid, increased mRNA gene expression was found at 7d for Atrogin-1/MAFbx and at 2d–7d for Myogenin. Both the volume and thickness of the masseter, temporalis, and medial pterygoid muscles from the BoNTA side were significantly reduced at 14d. In contrast, the lateral pterygoid from the BoNTA side showed a significant increase in volume at 14d. Therefore, the unilateral hypofunction of the masseter leads to molecular and morphological signs of atrophy in both the BoNTA-injected muscle and its agonistic non-injected masticatory muscles. The generalized effect on the mouse masticatory apparatus when one of its components is intervened suggests the need for more clinical studies to determine the safety of BoNTA usage in clinical dentistry.Item A Bisphosphonate With a Low Hydroxyapatite Binding Affinity Prevents Bone Loss in Mice After Ovariectomy and Reverses Rapidly With Treatment Cessation(Wiley, 2021-03-03) Coffman, Abigail A.; Basta-Pljakic, Jelena; Guerra, Rosa M.; Ebetino, Frank H.; Lundy, Mark W.; Majeska, Robert J.; Schaffler, Mitchell B.; Anatomy, Cell Biology and Physiology, School of MedicineBisphosphonates (BPs) are a mainstay of osteoporosis treatment; however, concerns about bone health based on oversuppression of remodeling remain. Long‐term bone remodeling suppression adversely affects bone material properties with microdamage accumulation and reduced fracture toughness in animals and increases in matrix mineralization and atypical femur fractures in patients. Although a “drug holiday” from BPs to restore remodeling and improve bone quality seems reasonable, clinical BPs have long functional half‐lives because of their high hydroxyapatite (HAP) binding affinities. This places a practical limit on the reversibility and effectiveness of a drug holiday. BPs with low HAP affinity and strong osteoclast inhibition potentially offer an alternative approach; their antiresorptive effect should reverse rapidly when dosing is discontinued. This study tested this concept using NE‐58025, a BP with low HAP affinity and moderate osteoclast inhibition potential. Young adult female C57Bl/6 mice were ovariectomized (OVX) and treated with NE‐58025, risedronate, or PBS vehicle for 3 months to test effectiveness in preventing long‐term bone loss. Bone microarchitecture, histomorphometry, and whole‐bone mechanical properties were assessed. To test reversibility, OVX mice were similarly treated for 3 months, treatment was stopped, and bone was assessed up to 3 months post‐treatment. NE‐58025 and RIS inhibited long‐term OVX‐induced bone loss, but NE‐58025 antiresorptive effects were more pronounced. Withdrawing NE‐58025 treatment led to the rapid onset of trabecular resorption with a 200% increase in osteoclast surface and bone loss within 1 month. Cessation of risedronate treatment did not lead to increases in resorption indices or bone loss. These results show that NE‐58025 prevents OVX‐induced bone loss, and its effects reverse quickly following cessation treatment in vivo. Low‐HAP affinity BPs may have use as reversible, antiresorptive agents with a rapid on/off profile, which may be useful for maintaining bone health with long‐term BP treatment.Item Evaluation of Tranexamic Acid and Calcium Chloride in Major Traumas in a Prehospital Setting: A Narrative Review(Wolters Kluwer, 2023) Bell, Kameron T.; Salmon, Chase M.; Purdy, Benjamin A.; Canfield, Scott G.; Anatomy, Cell Biology and Physiology, School of MedicineExcessive blood loss in the prehospital setting poses a significant challenge and is one of the leading causes of death in the United States. In response, emergency medical services (EMS) have increasingly adopted the use of tranexamic acid (TXA) and calcium chloride (CaCl2) as therapeutic interventions for hemorrhagic traumas. Tranexamic acid functions by inhibiting plasmin formation and restoring hemostatic balance, while calcium plays a pivotal role in the coagulation cascade, facilitating the conversion of factor X to factor Xa and prothrombin to thrombin. Despite the growing utilization of TXA and CaCl2 in both prehospital and hospital environments, a lack of literature exists regarding the comparative effectiveness of these agents in reducing hemorrhage and improving patient outcomes. Notably, Morgan County Indiana EMS recently integrated the administration of TXA with CaCl2 into their treatment protocols, offering a valuable opportunity to gather insight and formulate updated guidelines based on patient-centered outcomes. This narrative review aims to comprehensively evaluate the existing evidence concerning the administration of TXA and CaCl2 in the prehospital management of hemorrhages, while also incorporating and analyzing data derived from the co-administration of these medications within the practices of Morgan County EMS. This represents the inaugural description of the concurrent use of both TXA and CaCl2 to manage hemorrhages in the scientific literature.Item Neurotrophic factors in the physiology of motor neurons and their role in the pathobiology and therapeutic approach to amyotrophic lateral sclerosis(Frontiers Media, 2023-08-24) Stansberry, Wesley M.; Pierchala, Brian A.; Anatomy, Cell Biology and Physiology, School of MedicineThe discovery of the neurotrophins and their potent survival and trophic effects led to great enthusiasm about their therapeutic potential to rescue dying neurons in neurodegenerative diseases. The further discovery that brain-derived neurotrophic factor (BDNF), ciliary neurotrophic factor (CNTF) and glial cell line-derived neurotrophic factor (GDNF) had potent survival-promoting activity on motor neurons led to the proposal for their use in motor neuron diseases such as amyotrophic lateral sclerosis (ALS). In this review we synthesize the literature pertaining to the role of NGF, BDNF, CNTF and GDNF on the development and physiology of spinal motor neurons, as well as the preclinical studies that evaluated their potential for the treatment of ALS. Results from the clinical trials of these molecules will also be described and, with the aid of decades of hindsight, we will discuss what can reasonably be concluded and how this information can inform future clinical development of neurotrophic factors for ALS.Item Electrical stimulation of hindlimb skeletal muscle has beneficial effects on sublesional bone in a rat model of spinal cord injury(Elsevier, 2021) Zhao, Wei; Peng, Yuanzhen; Hu, Yizhong; Guo, X. Edward; Li, Jiliang; Cao, Jay; Pan, Jiangping; Feng, Jian Q.; Cardozo, Christopher; Jarvis, Jonathan; Bauman, William A.; Qin, Weiping; Anatomy, Cell Biology and Physiology, School of MedicineSpinal cord injury (SCI) results in marked atrophy of sublesional skeletal muscle and substantial loss of bone. In this study, the effects of prolonged electrical stimulation (ES) and/or testosterone enanthate (TE) on muscle mass and bone formation in a rat model of SCI were tested. Compared to sham-transected animals, a significant reduction of the mass of soleus, plantaris and extensor digitorum longus (EDL) muscles was observed in animals 6 weeks post-SCI. Notably, ES or ES + TE resulted in the increased mass of the EDL muscles. ES or ES + TE significantly decreased mRNA levels of muscle atrophy markers (e.g., MAFbx and MurF1) in the EDL. Significant decreases in bone mineral density (BMD) (-27%) and trabecular bone volume (-49.3%) at the distal femur were observed in animals 6 weeks post injury. TE, ES and ES + TE treatment significantly increased BMD by +6.4%, +5.4%, +8.5% and bone volume by +22.2%, and +56.2% and+ 60.2%, respectively. Notably, ES alone or ES + TE resulted in almost complete restoration of cortical stiffness estimated by finite element analysis in SCI animals. Osteoblastogenesis was evaluated by colony-forming unit-fibroblastic (CFU-F) staining using bone marrow mesenchymal stem cells obtained from the femur. SCI decreased the CFU-F+ cells by -56.8% compared to sham animals. TE or ES + TE treatment after SCI increased osteoblastogenesis by +74.6% and +67.2%, respectively. An osteoclastogenesis assay revealed significantly increased TRAP+ multinucleated cells (+34.8%) in SCI animals compared to sham animals. TE, ES and TE + ES treatment following SCI markedly decreased TRAP+ cells by -51.3%, -40.3% and -46.9%, respectively. Each intervention greatly reduced the ratio of RANKL to OPG mRNA of sublesional long bone. Collectively, our findings demonstrate that after neurologically complete paralysis, dynamic muscle resistance exercise by ES reduced muscle atrophy, downregulated genes involved in muscle wasting, and restored mechanical loading to sublesional bone to a degree that allowed for the preservation of bone by inhibition of bone resorption and/or by facilitating bone formation.Item Highly sensitive lipid detection and localization in atherosclerotic plaque with a dual-frequency intravascular photoacoustic/ultrasound catheter(Wiley, 2020) Cao, Yingchun; Alloosh, Mouhamad; Sturek, Michael; Cheng, Ji-Xin; Anatomy, Cell Biology and Physiology, School of MedicineIntravascular photoacoustic/ultrasound (IVPA/US) is an emerging hybrid imaging modality that provides specific lipid detection and localization, while maintaining co-registered artery morphology, for diagnosis of vulnerable plaque in cardiovascular disease. However, current IVPA/US approaches based on a single-element transducer exhibit compromised performance for lipid detection due to the relatively low contrast of lipid absorption and conflicting detection bands for photoacoustic and ultrasound signals. Here, we present a dual-frequency IVPA/US catheter for highly sensitive detection and precision localization of lipids. The low frequency transducer provides enhanced photoacoustic sensitivity, while the high frequency transducer maintains state-of-the-art spatial resolution for ultrasound imaging. The boosted capability of IVPA/US imaging enables a multi-scale analysis of lipid distribution in swine with coronary atherosclerosis. The dual-frequency IVPA/US catheter has a diameter of 1 mm and flexibility to easily adapt to current catheterization procedures and is a significant step toward clinical diagnosis of vulnerable plaque.Item Animal models for musculoskeletal research(Elsevier, 2023) Plotkin, Lilian I.; Kalajzic, Ivo; Anatomy, Cell Biology and Physiology, School of MedicineItem The Effect of SERCA Activation on Functional Characteristics and Signaling of Rat Soleus Muscle upon 7 Days of Unloading(MDPI, 2023-09-06) Sharlo, Kristina A.; Lvova, Irina D.; Tyganov, Sergey A.; Zaripova, Ksenia A.; Belova, Svetlana P.; Kostrominova, Tatiana Y.; Shenkman, Boris S.; Nemirovskaya, Tatiana L.; Anatomy, Cell Biology and Physiology, School of MedicineSkeletal muscle abnormalities and atrophy during unloading are accompanied by the accumulation of excess calcium in the sarcoplasm. We hypothesized that calcium accumulation may occur, among other mechanisms, due to the inhibition of sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) activity. Consequently, the use of the SERCA activator will reduce the level of calcium in the sarcoplasm and prevent the negative consequences of muscle unloading. Wistar rats were randomly assigned into one of three groups (eight rats per group): control rats with placebo (C), 7 days of unloading/hindlimb suspension with placebo (7HS), and 7 days of unloading treated with SERCA activator CDN1163 (7HSC). After seven days of unloading the soleus muscle, the 7HS group displayed increased fatigue in the ex vivo test, a significant increase in the level of calcium-dependent CaMK II phosphorylation and the level of tropomyosin oxidation, as well as a decrease in the content of mitochondrial DNA and protein, slow-type myosin mRNA, and the percentage of slow-type muscle fibers. All of these changes were prevented in the 7HSC group. Moreover, treatment with CDN1163 blocked a decrease in the phosphorylation of p70S6k, an increase in eEF2 phosphorylation, and an increase in MuRF-1 mRNA expression. Nevertheless, there were no differences in the degree of fast and slow muscle fiber atrophy between the 7HS and 7HSC groups. Conclusion: SERCA activation during 7 days of unloading prevented an increase in soleus fatigue, the decrease of slow-type myosin, mitochondrial markers, and markers of calcium homeostasis but had no effect on muscle atrophy.Item A crystallin mutant cataract with mineral deposits(Elsevier, 2023) Minogue, Peter J.; Gao, Junyuan; Mathias, Richard T.; Williams, James C., Jr.; Bledsoe, Sharon B.; Sommer, Andre J.; Beyer, Eric C.; Berthoud, Viviana M.; Anatomy, Cell Biology and Physiology, School of MedicineConnexin mutant mice develop cataracts containing calcium precipitates. To test whether pathologic mineralization is a general mechanism contributing to the disease, we characterized the lenses from a nonconnexin mutant mouse cataract model. By cosegregation of the phenotype with a satellite marker and genomic sequencing, we identified the mutant as a 5-bp duplication in the γC-crystallin gene (Crygcdup). Homozygous mice developed severe cataracts early, and heterozygous animals developed small cataracts later in life. Immunoblotting studies showed that the mutant lenses contained decreased levels of crystallins, connexin46, and connexin50 but increased levels of resident proteins of the nucleus, endoplasmic reticulum, and mitochondria. The reductions in fiber cell connexins were associated with a scarcity of gap junction punctae as detected by immunofluorescence and significant reductions in gap junction-mediated coupling between fiber cells in Crygcdup lenses. Particles that stained with the calcium deposit dye, Alizarin red, were abundant in the insoluble fraction from homozygous lenses but nearly absent in wild-type and heterozygous lens preparations. Whole-mount homozygous lenses were stained with Alizarin red in the cataract region. Mineralized material with a regional distribution similar to the cataract was detected in homozygous lenses (but not wild-type lenses) by micro-computed tomography. Attenuated total internal reflection Fourier-transform infrared microspectroscopy identified the mineral as apatite. These results are consistent with previous findings that loss of lens fiber cell gap junctional coupling leads to the formation of calcium precipitates. They also support the hypothesis that pathologic mineralization contributes to the formation of cataracts of different etiologies.