Mechanical stretch affects the healing and remodeling process of the anterior cruciate ligament (ACL) after surgery in important ways. In this study, the effects of mechanical stress on gene expression of type I and III collagen by cultured human ACL cells and roles of transforming growth factor (TGF)-beta1 in the regulation of mechanical strain-induced gene expression were investigated. Uniaxial cyclic stretch was applied on ACL cells at 10 cycles/min with 10% length stretch for 24 h. mRNA expression of the type I and type III collagen was increased by the cyclic stretch. TGF-beta1 protein in the cell culture supernatant was also increased by the stretch. In the presence of anti-TGF-beta1 antibody, stretch-induced increase in type I and type III mRNA expression was markedly ablated. The results suggest that the stretch-induced mRNA expression of the type I and type III collagen is mediated via an autocrine mechanism of TGF-beta1 released from ligament cells.

Maternal transmission of mitochondrial DNA (mtDNA) allows us to generate mtDNA congenic strain by repeating backcrosses of female mice to male mice of an inbred strain, which carries different mtDNA haplotype from that of the female progenitor. Since genetic backgrounds of inbred strains commonly used (e.g., C57BL/6J [B6] and BALB/c) are mainly derived from an European subspecies of Mus musculus domesticus, congenic strains, in which mtDNA originated from an Asian subspecies M. musculus musculus or an European species M. spretus, give in vivo condition that mismatch occurs between the mitochondrial and the nuclear genome. So far, little has been known how the mismatch condition affects the physiological phenotype of the mice. To address this question, we established two mtDNA congenic strains, C57BL/6J(B6)-mtSPR and BALB/c-mtSHH, which carry M. spretus- and M. m. musculus-derived mtDNAs, representing the conditions of interspecific and intersubspecific mitochondrial-nuclear genome mismatch, respectively. Using these congenic strains, we examined their physical performance by measuring their running time on a treadmill belt until exhaustion. The result clearly showed that the mtDNA congenic strains manifested a significant decrease in the level of physical performance, when compared with their progenitor strains. It also appeared that the congenic mice manifested growth rate. Thus, all results indicated that mismatch between the mitochondrial and the nuclear genome causes phenotypic changes in individuals of mice.

Local hyperthermia has been widely used as physical therapy for a number of diseases such as inflammatory osteoarticular disorders, tendinitis, and muscle injury. Local hyperthermia is clinically applied to improve blood and lymphatic flow to decrease swelling of tissues (e.g., skeletal muscle). As for muscle repair following injury, the mechanisms underlying the beneficial effects of hyperthermia-induced muscle repair are unknown. In this study, we investigated the direct effects of continuous heat stress on the differentiation of cultured mammalian myoblasts. Compared with control cultures grown at 37°C, incubation at 39°C (continuous mild heat stress; CMHS) enhanced myotube diameter, whereas myotubes were poorly formed at 41°C by primary human skeletal muscle culture cells, human skeletal muscle myoblasts (HSMMs), and C2C12 mouse myoblasts. In HSMMs and C2C12 cells exposed to CMHS, mRNA and protein levels of myosin heavy chain (MyHC) type I were increased compared with the control cultures. The mRNA level of MyHC IIx was unaltered in HSMMs and decreased in C2C12 cells, compared with cells that were not exposed to heat stress. These results indicated a fast-to-slow fiber-type shift in myoblasts. We also examined upstream signals that might be responsible for the fast-to-slow shift of fiber types. CMHS enhanced the mRNA and protein levels of peroxisome proliferator-activated receptor-γ coactivator (PGC)-1α in HSMMS and C2C12 cells but not the activities of MAPKs (ERK1/2 and p38 MAPK) in HSMMs and C2C12 cells. These data suggest that CMHS induces a fast-to-slow fiber-type shift of mammalian myoblasts through PGC-1α.

The effects of ovarian hormones on glucose and fatty acid oxidation during exercise were investigated in adult female ovariectomized rats. Rats subdivided into 3 groups received intraperitoneal injections of hormones or sesame oil for 8 days. Estrogen (E) treated rats received 17-β estradiol in daily doses of 2 μg. Estrogen and progesterone treated rats (EP) received 17-β estradiol in daily doses of 2 μg and 2 mg, respectively. Control rats (S) received sesame oil alone. After an overnight fast, rats ran at the speed of 25 m · min^-1 for 60 min. [U-¹⁴C]glucose or [1-¹⁴C]palmitate was injected into rats at 5 min of exercise and before 10 min of exercise, respectively. Expired ¹⁴CO₂ was collected using bottomless chamber on a treadmill belt. No significant differences were found in mean blood glucose, lactate and plasma free fatty acid concentrations after the exercise. Until the end of the exercise 34.7±2.6 (E, n=5), 40.8±2.9 (EP, n=5) and 43.7± 3.5% (S, n=6) (mean±SE) of ¹⁴C which was injected as ¹⁴C-glucose was recovered as ¹⁴CO₂. During 60 min of the exercise 27.5±1.0 (E, n=7), 19.8±2.7 (EP, n=6) and 25.0±1.9% (S, n=6) of ¹⁴C which was injected as ¹⁴C-palmitate was recovered as ¹⁴CO₂. A significant difference was found in this rate between E and EP (P < 0.05). It was concluded that estrogen treatment stimulated fatty acid oxidation compared with the estrogen plus progesterone treatment and tended to inhibit glucose oxidation during prolonged exercise.

Atrophy of rat soleus muscles by hindlimb suspension is characterized by an early dramatic decrease in a soluble 22-kDa protein. The 22-kDa protein was purified from rat red skeletal muscle and rat lens by three different methods of chromatography. The partial amino acid sequence (66% of total amino acids) determined for muscle 22-kDa protein was identical with that of rat lens crystallin. The HPLC elution patterns of lysylendopeptidase fragments of 22-kDa protein from the two sources were identical. Polyclonal antibodies to rat muscle and bovine lens αB-crystallin with the two proteins on immunoblotting. αB-Crystallin protein was expressed and synthesized efficiently in slow skeletal muscle and poorly in fast muscle. Thus, the decreased 22-kDa protein of slow muscle in the suspension treatment was confirmed to be αB-crystallin. Immunoblotting confirmed that most of the αB-crystallin was solubilized, though some was tightly bound to myofibrils. This bound portion was localized in Z-bands of isolated myofibrils by immunocyto-chemical light and electron microscopy. Muscle αB-crystallin is tentatively proposed to be a myofibril-stabilizing protein, based upon its extraction characteristics, localization, and amino acid sequence.

αB-Crystallin, one of the small heat shock proteins, is constitutively expressed in lens as well as in nonlenticular tissues. It can function as a molecular chaperone for other lens crystallins and some other proteins. Its nonocular function is unknown although some reported one of them is related to cytoskeletal networks and/or components. In the present study, we demonstrate the association of αB-crystallin with tubulin. Imnninoprecipitation experiments using L6 myoblast cell lysate with anti-αB-crystallin antibody resulted in the coprecipitation of α-tubulin, which was apparently temperature-dependent. Further, purified αB-crystallin prevented the turbidity development of purified tubulin molecule at 37°C in vitro. Sucrose gradient centrifugation revealed that this chaperone activity was accompanied by the formation of large complex of αB-crystallin and tubulin dimer. These results indicate that one of the nonlenticular functions of αB-crystallin may be the protection of tubulin subunits of microtubules.