Muscle Cell Nuclei Research Articles

Effects of photobiomodulation therapy and topical non-steroidal anti-inflammatory drug on skeletal muscle injury induced by contusion in rats-part 1: morphological and functional aspects.

Musculoskeletal injuries are very frequent and are responsible for causing pain and impairment of muscle function, as well as significant functional limitations. In the acute phase, the most prescribed treatment is with non-steroidal anti-inflammatory drugs (NSAIDs), despite their questionable effectiveness. However, the use of photobiomodulation therapy (PBMT) in musculoskeletal disorders has been increasing in the last few years, and this therapy appears to be an interesting alternative to the traditional drugs. The objective of the present study was to evaluate and compare the effects of PBMT, with different application doses, and topical NSAIDs, under morphological and functional parameters, during an acute inflammatory process triggered by a controlled model of musculoskeletal injury induced via contusion in rats. Muscle injury was induced by means of a single trauma to the animals' anterior tibialis muscle. After 1h, the rats were treated with PBMT (830nm; continuous mode, with a power output of 100mW; 3.57W/cm2; 1J-35.7J/cm2, 3J-107.1J/cm2, and 9J-321.4J/cm2; 10, 30, and 90s) or diclofenac sodium for topical use (1g). Morphological analysis (histology) and functional analysis (muscle work) were performed, 6, 12, and 24h after induction of the injury. PBMT, with all doses tested, improved morphological changes caused by trauma; however, the 9J (321.4J/cm2) dose was the most effective in organizing muscle fibers and cell nuclei. On the other hand, the use of diclofenac sodium produced only a slight improvement in morphological changes. Moreover, we observed a statistically significant increase of muscle work in the PBMT 3J (107.1J/cm2) group in relation to the injury group and the diclofenac group (p<0.05). The results of the present study indicate that PBMT, with a dose of 3J (107.1J/cm2), is more effective than the other doses of PBMT tested and NSAIDs for topical use as a means to improve morphological and functional alterations due to muscle injury from contusion.

Chloride intracellular channel proteins respond to heat stress in Caenorhabditis elegans.

Chloride intracellular channel proteins (CLICs) are multi-functional proteins that are expressed in various cell types and differ in their subcellular location. Two CLIC homologs, EXL-1 (excretory canal abnormal like-1) and EXC-4 (excretory canal abnormal– 4), are encoded in the Caenorhabditis elegans genome, providing an excellent model to study the functional diversification of CLIC proteins. EXC-4 functions in excretory canal formation during normal animal development. However, to date, the physiological function of EXL-1 remains largely unknown. In this study, we demonstrate that EXL-1 responds specifically to heat stress and translocates from the cytoplasm to the nucleus in intestinal cells and body wall muscle cells under heat shock. In contrast, we do not observe EXC-4 nuclear translocation under heat shock. Full protein sequence analysis shows that EXL-1 bears a non-classic nuclear localization signal (NLS) that EXC-4 is lacking. All mammalian CLIC members have a nuclear localization signal, with the exception of CLIC3. Our phylogenetic analysis of the CLIC gene families across various animal species demonstrates that the duplication of CLICs in protostomes and deuterostomes occurred independently and that the NLS was subsequently lost in amniotes and nematodes, suggesting convergent evolution. We also observe that EXL-1 nuclear translocation occurs in a timely ordered manner in the intestine, from posterior to anterior regions. Finally, we find that exl-1 loss of function mutants are more susceptible to heat stress than wild-type animals, demonstrating functional relevance of the nuclear translocation. This research provides the first link between CLICs and environmental heat stress. We propose that C. elegans CLICs evolved to achieve different physiological functions through subcellular localization change and spatial separation in response to external or internal signals.

Histopathologic differences partially distinguish syndromic aortic diseases.

A variety of syndromic diseases such as Marfan syndrome, Loeys-Dietz syndrome, and bicuspid aortic valve with aneurysm along with risk factors of smoking and hypertension result in ascending aortic aneurysms and dissections. Historically, a complicated variety of terms have been used to describe a range of histopathologies that are present in resected specimens. As a result, no consistent patterns of histopathology have been reported. We used the recent Society for Cardiovascular Pathology/Association for European Cardiovascular Pathology consensus statement on nomenclature and diagnostic criteria for noninflammatory aortic disease to blindly evaluate 148 surgically resected specimens. We found that overall patterns of histopathologic changes could separately cluster bicuspid aortic valve and nonsyndromic subjects from Marfan and Loeys-Dietz subjects. Marfan syndrome cases significantly had more overall medial degeneration and mucoid extracellular matrix accumulation than other syndromes. Smooth muscle cell nuclei loss was a feature of aging and not a feature of Marfan or Loeys-Dietz syndrome subjects. We conclude that a consistent use of histologic and histopathologic descriptors can help discriminate different etiologies of ascending aortic aneurysms.

Cell membrane integrity and revascularization: The possible functional mechanism of ischemic preconditioning for skeletal muscle protection against ischemic-reperfusion injury

BackgroundThe purpose of this paper was to evaluate whether ischemic preconditioning (IPC) could make protective effects against skeletal muscle injuries induced by ischemic-reperfusion (I/R). MethodsEighteen rats were randomly divided into three groups of 6 subjects each: control group, I/R group, and IPC group. Thigh root ischemia of rats in the I/R group was induced by 3h ischemia and 24h reperfusion. IPC was applied by 3 periods of 15min ischemia/15min reperfusion prior to ischemia. Morphological changes in skeletal muscle cells induced by I/R and IPC were observed by hematoxylin and eosin (HE) staining and electron microscopy. In addition, angiogenesis was evaluated by immunolabeling of CD31. ResultsIPC could prevented morphological alternations induced by ischemia, including myofilament, cell membrane, cell matrix, nucleus, mitochondria, and sarcoplasmic reticulum damage in skeletal muscle cells. The CD31 immunolabeling showed that neovascularization was observed in the IPC group but not in the I/R group. IPC could protect skeletal muscle cells from necrosis, apoptosis, and morphological damages induced by I/R injury. ConclusionRevascularization may play a key role in the mechanism underlying the protective effects of IPC in vivo.

Calcium homeostasis alterations in a mouse model of the Dynamin 2-related centronuclear myopathy.

ABSTRACTAutosomal dominant centronuclear myopathy (CNM) is a rare congenital myopathy characterized by centrally located nuclei in muscle fibers. CNM results from mutations in the gene encoding dynamin 2 (DNM2), a large GTPase involved in endocytosis, intracellular membrane trafficking, and cytoskeleton regulation. We developed a knock-in mouse model expressing the most frequent DNM2-CNM mutation; i.e. the KI-Dnm2R465W model. Heterozygous (HTZ) KI-Dnm2 mice progressively develop muscle atrophy, impairment of contractile properties, histopathological abnormalities, and elevated cytosolic calcium concentration. Here, we aim at better characterizing the calcium homeostasis impairment in extensor digitorum longus (EDL) and soleus muscles from adult HTZ KI-Dnm2 mice. We demonstrate abnormal contractile properties and cytosolic Ca2+ concentration in EDL but not soleus muscles showing that calcium impairment is correlated with muscle weakness and might be a determinant factor of the spatial muscle involvement. In addition, the elevated cytosolic Ca2+ concentration in EDL muscles is associated with an increased sarcolemmal permeability to Ca2+ and releasable Ca2+ content from the sarcoplasmic reticulum. However, amplitude and kinetics characteristics of the calcium transient appear unchanged. This suggests that calcium defect is probably not a primary cause of decreased force generation by compromised sarcomere shortening but may be involved in long-term deleterious consequences on muscle physiology. Our results highlight the first pathomechanism which may explain the spatial muscle involvement occurring in DNM2-related CNM and open the way toward development of a therapeutic approach to normalize calcium content.

The ecdysone receptor (EcR) is a major regulator of tissue development and growth in the marine salmonid ectoparasite, Lepeophtheirus salmonis (Copepoda, Caligidae)

The function of the ecdysone receptor (EcR) during development and molting has been thoroughly investigated in some arthropods such as insects but rarely in crustacean copepods such as the salmon louse Lepeophtheirus salmonis (L. salmonis) (Copepoda, Caligidae). The salmon louse is an ectoparasite on Atlantic salmon that has major economical impact in aquaculture due to the cost of medical treatment methods to remove lice from the fish. Handling of salmon louse infestations is further complicated by development of resistance towards available medicines. Understanding of basic molecular biological processes in the salmon louse is essential to enable development of new tools to control the parasite. In this study, we found L. salmonis EcR (LsEcR) transcript to be present in the neuronal somata of the brain, nuclei of muscle fibres and the immature intestine of the salmon louse. Furthermore, we explored the function of LsEcR during development using RNA interference mediated knock-down and through infection trials. Our results show that knock-down of LsEcR in the salmon louse is associated with hypotrophy of several tissues, delayed development and mortality. In addition, combined knock-down of LsEcR/LsRXR resulted in molting arrest during early larval stages.

Phenylbutazone induces expression of MBNL1 and suppresses formation of MBNL1-CUG RNA foci in a mouse model of myotonic dystrophy.

Myotonic dystrophy type 1 (DM1) is caused by abnormal expansion of CTG repeats in the 3′ untranslated region of the DMPK gene. Expanded CTG repeats are transcribed into RNA and make an aggregate with a splicing regulator, MBNL1, in the nucleus, which is called the nuclear foci. The nuclear foci sequestrates and downregulates availability of MBNL1. Symptomatic treatments are available for DM1, but no rational therapy is available. In this study, we found that a nonsteroidal anti-inflammatory drug (NSAID), phenylbutazone (PBZ), upregulated the expression of MBNL1 in C2C12 myoblasts as well as in the HSALR mouse model for DM1. In the DM1 mice model, PBZ ameliorated aberrant splicing of Clcn1, Nfix, and Rpn2. PBZ increased expression of skeletal muscle chloride channel, decreased abnormal central nuclei of muscle fibers, and improved wheel-running activity in HSALR mice. We found that the effect of PBZ was conferred by two distinct mechanisms. First, PBZ suppressed methylation of an enhancer region in Mbnl1 intron 1, and enhanced transcription of Mbnl1 mRNA. Second, PBZ attenuated binding of MBNL1 to abnormally expanded CUG repeats in cellulo and in vitro. Our studies suggest that PBZ is a potent therapeutic agent for DM1 that upregulates availability of MBNL1.

Consensus statement on surgical pathology of the aorta from the Society for Cardiovascular Pathology and the Association For European Cardiovascular Pathology: II. Noninflammatory degenerative diseases - nomenclature and diagnostic criteria.

Surgical aortic specimens are usually examined in Pathology Departments as a result of treatment of aneurysms or dissections. A number of diseases, genetic syndromes (Marfan syndrome, Loeys-Dietz syndrome, etc.), and vasculopathic aging processes involved in vascular injury can cause both distinct and nonspecific histopathologic changes with degeneration of the media as a common denominator. Terminology for these changes has varied over time leading to confusion and inconsistencies. This consensus document has established a revised, unified nomenclature for the variety of noninflammatory degenerative aortic histopathologies seen in such specimens. Older terms such as cystic medial necrosis and medionecrosis are replaced by more technically accurate terms such as mucoid extracellular matrix accumulation (MEMA), elastic fiber fragmentation and/or loss, and smooth muscle cell nuclei loss. A straightforward system of grading is presented to gauge the extent of medial degeneration and synoptic reporting tables are provided. Herein we present a standardized nomenclature that is accessible to general pathologists and useful for future publications describing these entities.

Molecular hydrogen alleviates motor deficits and muscle degeneration in mdx mice

Objective: Duchenne muscular dystrophy (DMD) is a devastating muscle disease caused by a mutation in DMD encoding dystrophin. Oxidative stress accounts for dystrophic muscle pathologies in DMD. We examined the effects of molecular hydrogen in mdx mice, a model animal for DMD.Methods: The pregnant mother started to take supersaturated hydrogen water (>5 ppm) ad libitum from E15.5 up to weaning of the offspring. The mdx mice took supersaturated hydrogen water from weaning until age 10 or 24 weeks when they were sacrificed.Results: Hydrogen water prevented abnormal body mass gain that is commonly observed in mdx mice. Hydrogen improved the spontaneous running distance that was estimated by a counter-equipped running-wheel, and extended the duration on the rota-rod. Plasma creatine kinase activities were decreased by hydrogen at ages 10 and 24 weeks. Hydrogen also decreased the number of central nuclei of muscle fibers at ages 10 and 24 weeks, and immunostaining for nitrotyrosine in gastrocnemius muscle at age 24 weeks. Additionally, hydrogen tended to increase protein expressions of antioxidant glutathione peroxidase 1, as well as anti-apoptotic Bcl-2, in skeletal muscle at age 10 weeks.Discussion: Although molecular mechanisms of the diverse effects of hydrogen remain to be elucidated, hydrogen potentially improves muscular dystrophy in DMD patients.

Особливості просторової і структурної організації шлунка білих лабораторних щурів у нормі

&lt;p&gt;Резюме. Дослідження проведено на 12 інтактних білих лабораторних щурах. Макрометрично вимірювали довжину, ширину і товщину шлунка з наступним обчисленням його об’єму. Для гістологічного дослідження забирали шматочки тканин із різних відділів шлунка. Морфометричні дослідження включали визначення товщини слизової, м’язової, серозної оболонок та підслизової основи, площі епітеліоцитів, діаметра ядер епітеліоцитів, ядерно-цитоплазматичних співвідношень (ЯЦС). Середні розміри шлунка у інтактних тварин: довжина – (3,93±0,13) см, ширина – (2,20±0,09) см, товщина – (1,45±0,08) см, об’єм шлунка у середньому складає (6,72±0,81) см³. Найбільший питомий об’єм має дно шлунка. При зовнішньому макроскопічному огляді дно шлунка за його білуватим забарвленням легко можна відрізнити від червоно-сіруватих тіла та пілоричної частини. Ще чіткіше дно шлунка можна віддиференціювати при візуальному огляді його слизової оболонки: слизова оболонка дна відмежована від тіла шлунка чіткою дугоподібною складкою або складчастим краєм. При гістологічному дослідженні складчастого краю встановлено, що його основу складає дуплікатра підслизової основи, яка зі сторони дна шлунка покрита багатошаровим плоским епітелієм, а зі сторони тіла – залозистим епітелієм. Морфометричні дослідження стінки шлунка інтактних білих лабораторних щурів дозволили встановити наступні кількісні параметри її структурних компонентів. Товщина серозної оболонки у середньому складала (7,83±0,31) мкм, м’язової – (40,17±0,75) мкм, підслизової – (24,00±0,78) мкм і слизової – (96,00±0,97) мкм. Площа ядер гладком’язових клітин (ГМК) дещо більша, ніж площа ядер залозистого епітелію, водночас питомий об’єм цитоплазми у ГМК, навпаки, менший. Відповідно до цього і показник ЯЦС у ГМК вищий, ніж у епітеліоцитів слизової.&lt;/p&gt;&lt;p&gt; &lt;/p&gt;&lt;p&gt;Ключові слова: шлунок щура, будова, морфометрія.&lt;/p&gt;

The zebrafish fast myosin light chain mylpfa:H2B-GFP transgene is a useful tool for in vivo imaging of myocyte fusion in the vertebrate embryo

BackgroundSkeletal muscle fibers are multinucleated syncytia that arise from the fusion of mononucleated precursors, the myocytes, during embryonic development, muscle hypertrophy in post-embryonic growth and muscle regeneration after injury. Even though myocyte fusion is central to skeletal muscle differentiation, our current knowledge of the molecular mechanism of myocyte fusion in the vertebrates is rather limited. Previous work, from our group and others, has shown that the zebrafish embryo is a very useful model for investigating the cell biology and genetics of vertebrate myocyte fusion in vivo. ResultsHere, we report the generation of a stable transgenic zebrafish strain that expresses the Histone 2B-GFP (H2B-GFP) fusion protein in the nuclei of all fast-twitch muscle fibers under the control of the fast-twitch muscle-specific myosin light chain, phosphorylatable, fast skeletal muscle a (mylpfa) gene promoter. By introducing this transgene into a mutant for junctional adhesion molecule 3b (jam3b), which encodes a cell adhesion protein previously implicated in myocyte fusion, we demonstrate the feasibility of using this transgene for the analysis of myocyte fusion during the differentiation of the trunk musculature of the zebrafish embryo. ConclusionsSince we know so little about the molecules regulating vertebrate myocyte fusion, we propose that the mylpfa:H2B-GFP transgene will be a very useful reporter for conducting forward and reverse genetic screens to identify new components regulating vertebrate myocyte fusion.

Activation of the wnt/β-Catenin Signaling Pathway in Polymyositis, Dermatomyositis and Duchenne Muscular Dystrophy.

Background and PurposeThe wnt/β-catenin signaling pathway plays a critical role in embryonic development and adult-tissue homeostasis. Recent investigations implicate the importance of wnt/β-catenin signaling in normal wound healing and its sustained activation being associated with fibrogenesis. We investigated the immunolocalization and activation of wnt/β-catenin in polymyositis (PM), dermatomyositis (DM), and Duchenne muscular dystrophy (DMD).MethodsImmunofluorescence staining and Western blot analysis of β-catenin were performed in muscle specimens from 6 PM, 8 DM, and 6 DMD subjects. The β-catenin/Tcf4 DNA-binding activity in muscle was studied using an electrophoretic mobility shift assay (EMSA), and serum wnt/β-catenin/Tcf transcriptional activity was measured using a luciferase reporter gene assay.ResultsImmunoreactivity for β-catenin was found in the cytoplasm and nuclei of muscle fibers in PM, DM, and DMD. The protein level of β-catenin was elevated, and EMSA analysis confirmed the activation of wnt/β-catenin signaling. The transcriptional activities of β-catenin/Tcf in the circulation were increased in patients with PM, DM, and DMD, especially in those with interstitial lung disease, and these transcriptional activities decreased when PM or DM patients exhibited obvious clinical improvements.ConclusionsOur findings indicate that wnt/β-catenin signaling is activated in PM, DM, and DMD. Its activation in muscle tissue and the circulation may play a role in modulating muscle regeneration and be at least partly involved in the process of muscle and pulmonary fibrosis.

Hemodynamic properties and arterial structure in male rat offspring with fetal hypothyroidism.

Thyroid hormones (THs) play a crucial role in the development of different systems during fetal life; fetal hypothyroidism (FH) is associated with reduced cardiac function and dimensions in neonates. The aim of this study is to determine whether TH deficiency during fetal life is associated with arterial structural and hemodynamic changes during adulthood. Hypothyroidism was induced by adding 0.025% 6-propyl-2-thiouracil in drinking water throughout pregnancy, while controls consumed only tap water. Hemodynamic parameters, cross-sectional area, intima-media thickness (IMT), and density of nuclei of smooth muscle cells and endothelial cells (ECs) in the aorta and mesenteric arteries were measured. Compared to controls, in the FH group, baseline systolic blood pressure (105.7 ± 3.1 vs. 87.9 ± 3.3 mm Hg, p < 0.01), diastolic blood pressure (64.4 ± 1.7 vs. 53.2 ± 2.1 mm Hg, p < 0.05), and mean arterial pressure (80.9 ± 2.1 vs. 67.1 ± 2.1 mm Hg, p < 0.01) were significantly lower. In addition, in the FH group, intensity and latency of response to phenylephrine were significantly lower and longer, respectively, as were the IMT and density of ECs in the aorta and superior mesenteric arteries. In conclusion, this study showed that TH deficiency during fetal life can have long-lasting functional and histological effects, which can compromise cardiovascular function during adulthood.

Immunoexpression of Steroid Hormone Receptors and Proliferation Markers in Uterine Leiomyoma and Normal Myometrial Tissues from the Miniature Pig, Sus scrofa.

Uterine leiomyomas in miniature pet pigs occur similarly to those in women with regard to frequency, age, parity, and cycling. Clinical signs, gross, and histologic features of the porcine tumors closely resemble uterine leiomyomas (fibroids) in women. Although fibroids are hormonally responsive in women, the roles of estrogen and progesterone have not been fully elucidated. In this study, immunohistochemistry was used to assess the expression of the steroid hormone receptors, estrogen receptor alpha (ER-α), estrogen receptor beta (ER-β) and progesterone receptor (PR), and cell proliferation markers, proliferating cell nuclear antigen (PCNA) and Ki-67 in tumor and matched myometrial tissues sampled from miniature pigs. A "quickscore" method was used to determine receptor expression and labeling indices were calculated for the markers. ER-α/β and PR were localized to the nuclei of smooth muscle cells in both tissues. PR expression was intense and diffuse throughout all tissues, with correlation between tumors and matched myometria. Conversely, ER-α expression was variable between the myometrial and tumor tissues, as well as between animals. ER-β expression was low. PCNA and Ki-67 were localized to the nucleus and expression varied among tumors; however, normal tissues were overall negative. These findings support further investigation into the use of the miniature pig as a model of fibroids in women.

High level of GHR nuclear translocation in skeletal muscle of a hyperplasic transgenic zebrafish.

It has been reported that nuclear translocation of growth hormone receptor (GHR) may directly activate cell proliferation in mammals and birds. However, this phenomenon has not yet been described in fish. Recently, we have developed a transgenic zebrafish that overexpresses GHR in a muscle-specific manner. Considering that this transgenic model exhibits hyperplasic muscle growth, the present work aims at verifying the relationship between GHR nuclear translocation and muscle cell proliferation. This relationship was evaluated by the phosphorylation state of the proliferative MEK/ERK pathway, expression of nuclear import-related genes, immunostaining of phospho-histone H3 (PH3) as a proliferation marker, and nuclear GHR localization. The results showed a significant decrease in the phosphorylation state of ERK1/2 proteins in transgenics. Moreover, there was an increase in expression of three out of four importin genes analyzed parallel to a large flow of GHR displacement toward and into the nucleus of transgenic muscle cells. Also, transgenics presented a marked increase in PH3 staining, which indicates cell proliferation. These findings, as far as we know, are the first report suggesting a proliferative action of GHR in fish as a consequence of its increased nuclear translocation. Thus, it appears that the nuclear migration of cytokine receptors is a common event among different taxonomic groups. In addition, the results presented here highlight the possibility that these membrane proteins may be involved more directly than previously thought in the control of genes related to cell growth and proliferation.

Localized nuclear and perinuclear Ca(2+) signals in intact mouse skeletal muscle fibers.

Nuclear Ca2+ is important for the regulation of several nuclear processes such as gene expression. Localized Ca2+ signals (LCSs) in skeletal muscle fibers of mice have been mainly studied as Ca2+ release events from the sarcoplasmic reticulum. Their location with regard to cell nuclei has not been investigated. Our study is based on the hypothesis that LCSs associated with nuclei are present in skeletal muscle fibers of adult mice. Therefore, we carried out experiments addressing this question and we found novel Ca2+ signals associated with nuclei of skeletal muscle fibers (with possibly attached satellite cells). We measured localized nuclear and perinuclear Ca2+ signals (NLCSs and PLCSs) alongside cytosolic localized Ca2+ signals (CLCSs) during a hypertonic treatment. We also observed NLCSs under isotonic conditions. The NLCSs and PLCSs are Ca2+ signals in the range of micrometer [FWHM (full width at half maximum): 2.75 ± 0.27 μm (NLCSs) and 2.55 ± 0.17 μm (PLCSs), S.E.M.]. Additionally, global nuclear Ca2+ signals (NGCSs) were observed. To investigate which type of Ca2+ channels contribute to the Ca2+ signals associated with nuclei in skeletal muscle fibers, we performed measurements with the RyR blocker dantrolene, the DHPR blocker nifedipine or the IP3R blocker Xestospongin C. We observed Ca2+ signals associated with nuclei in the presence of each blocker. Nifedipine and dantrolene had an inhibitory effect on the fraction of fibers with PLCSs. The situation for the fraction of fibers with NLCSs is more complex indicating that RyR is less important for the generation of NLCSs compared to the generation of PLCSs. The fraction of fibers with NLCSs and PLCSs is not reduced in the presence of Xestospongin C. The localized perinuclear and intranuclear Ca2+ signals may be a powerful tool for the cell to regulate adaptive processes as gene expression. The intranuclear Ca2+ signals may be particularly interesting in this respect.

Immunohistochemical characteristic of myoma tissue in patients with uterine leiomyoma after treatment with ulipristal acetate.

Background. Uterine leiomyoma is one of the most common benign tumors of the female genital organs. The main conservative treatment of leiomyoma is progesterone receptor blockers that suppress myoma growth and may lead to its regression. Objective. To study the immunohistochemical features of myoma tissue in patients with uterine leiomyoma after treatment with selective progesterone modulator - ulipristal acetate. Methods. Leiomyoma tissue obtained from 9 patients after ulipristal acetate treatment were investigated. Group for comparison - leiomyoma from patients without hormonal therapy. Immunohistochemical study of progesterone and estrogen receptors, proliferative activity marker Ki-67 and inhibitor of apoptosis Bcl-2 was performed. Results. In the group of patients without preoperative hormonal treatment progesterone receptors were expressed in 76,4±6,8% of the nuclei, estrogen receptors - in 32,8±2,6%. In the group of patients after treatment with ulipristal acetate there was a significant decrease of progesterone receptor expression – 36,8±1,28% (p &lt;0,05) and a nonsignificant decrease of estrogen expression – 30,7±3,4% (p&gt; 0,05) . Bcl-2 in the control group was found in 65,4±7,2% cells, in leiomyoma after treatment there was a significant decrease of bcl-2 – 42,6±3,2% (p &lt;0, 05). In leiomyomas without hormonal treatment Ki-67 was determined in 11,8% of the nuclei of smooth muscle cells, and in leiomyomas after ulipristal acetate – in 7,2% leiomyoma cells. Conclusions. In patients after three months of ulipristal acetate treatment there was a significant decrease of expression of progesterone receptor, bcl-2, and Ki-67. Taken together these data evidence reduced action of progesterone on leiomyoma cells, induction of apoptosis and decreased proliferation processes that may cause involution of fibroids.

Effects of supplemental lysine and methionine with zilpaterol hydrochloride on feedlot performance, carcass merit, and skeletal muscle fiber characteristics in finishing feedlot cattle.

Feeding zilpaterol hydrochloride (ZH) with ruminally protected AA was evaluated in a small-pen feeding trial. Crossbred steers ( = 180; initial BW = 366 kg) were blocked by weight and then randomly assigned to treatments (45 pens; 9 pens/treatment). Treatment groups consisted of no ZH and no AA (Cont-), ZH and no AA (Cont+), ZH and a ruminally protected lysine supplement (Lys), ZH and a ruminally protected methionine supplement (Met), and ZH and ruminally protected lysine and methionine (Lys+Met). Zilpaterol hydrochloride (8.3 mg/kg DM) was fed for the last 20 d of the finishing period with a 3-d withdrawal period. Lysine and Met were top dressed daily for the 134-d feeding trial to provide 12 or 4 g·hd·d, respectively, to the small intestine. Carcass characteristics, striploins, and prerigor muscle samples were collected following harvest at a commercial facility. Steaks from each steer were aged for 7, 14, 21, and 28 d, and Warner-Bratzler shear force (WBSF) was determined as an indicator of tenderness. Prerigor muscle samples were used for immunohistological analysis. Cattle treated with Met and Lys+Met had increased final BW ( < 0.3) and ADG ( < 0.05) compared to Cont- and Cont+. Supplementation of Lys, Met, and Lys+Met improved G:F ( < 0.05) compared to Cont- during the ZH feeding period (d 111 to 134) as well as the entire feeding period ( < 0.05). Zilpaterol hydrochloride increased carcass ADG ( < 0.05) when compared to non-ZH-fed steers. Methionine and Lys+Met treatments had heavier HCW ( < 0.02) than that of Cont-. Yield grade was decreased ( < 0.04) for Cont+ steers compared to steers treated with Lys, Lys+Met, and Cont-. Tenderness was reduced ( < 0.05) with ZH regardless of AA supplementation. Lysine, Met, Lys+Met, and Cont+ had less tender steaks ( < 0.05) throughout all aging groups compared to Cont-. Steaks from Lys-treated steers were less tender ( < 0.05) than those of Cont+ during the 7- and 14-d aging periods. Nuclei density was the greatest with Cont- cattle compared to all other treatments suggesting a dilution effect of the nuclei in the larger muscle fibers with ZH feeding. Supplementation of Met in conjunction with ZH feeding increased ADG and HCW although this may lead to decreased tenderness even after aging for 28 d. These findings indicated that steers fed ZH may require additional AA absorbed from the small intestine to maximize performance.

Versican localizes to the nucleus in proliferating mesenchymal cells.

Versican is a versatile and highly interactive chondroitin sulfate proteoglycan that is found in the extracellular matrix (ECM) of many tissues and is a major component of developing and developed lesions in atherosclerotic vascular disease. In this paper, we present data to indicate that versican may have important intracellular functions in addition to its better known roles in the ECM. Rat aortic smooth muscle cells were fixed and immunostained for versican and images of fluorescently labeled cells were obtained by confocal microscopy. Intracellular versican was detected in the nucleus and cytosol of vascular smooth muscle cells. The use of a synthetic neutralizing peptide eliminated versican immunostaining, demonstrating the specificity of the antibody used in this study. Western blot of pure nuclear extracts confirmed the presence of versican in the nucleus, and multifluorescent immunostaining showed strong colocalization of versican and nucleolin, suggesting a nucleolar localization of versican in nondividing cells. In dividing valve interstitial cells, a strong signal for versican was observed in and around the condensed chromosomes during the various stages of mitosis. Multifluorescent immunostaining for versican and tubulin revealed versican aggregated at opposing poles of the mitotic spindle during metaphase. Knockdown of versican expression using siRNA disrupted the organization of the mitotic spindle and led to the formation of multipolar spindles during metaphase. Collectively, these data suggest an intracellular function for versican in vascular cells where it appears to play a role in mitotic spindle organization during cell division. These observations open a new avenue for studies of versican, suggesting even more diverse roles in vascular health and disease.

Troponin T3 regulates nuclear localization of the calcium channel Cavβ1a subunit in skeletal muscle

The voltage-gated calcium channel (Cav) β1a subunit (Cavβ1a) plays an important role in excitation–contraction coupling (ECC), a process in the myoplasm that leads to muscle-force generation. Recently, we discovered that the Cavβ1a subunit travels to the nucleus of skeletal muscle cells where it helps to regulate gene transcription. To determine how it travels to the nucleus, we performed a yeast two-hybrid screening of the mouse fast skeletal muscle cDNA library and identified an interaction with troponin T3 (TnT3), which we subsequently confirmed by co-immunoprecipitation and co-localization assays in mouse skeletal muscle in vivo and in cultured C2C12 muscle cells. Interacting domains were mapped to the leucine zipper domain in TnT3 COOH-terminus (160–244 aa) and Cavβ1a NH2-terminus (1–99 aa), respectively. The double fluorescence assay in C2C12 cells co-expressing TnT3/DsRed and Cavβ1a/YFP shows that TnT3 facilitates Cavβ1a nuclear recruitment, suggesting that the two proteins play a heretofore unknown role during early muscle differentiation in addition to their classical role in ECC regulation.