Functional and molecular outcomes of the human masticatory muscles.

The masticatory muscles achieve a broad range of different activities such as chewing, sucking, swallowing, and speech. In order to accomplish these duties, masticatory muscles have a unique and heterogeneous structure and fiber composition, enabling them to produce their strength and contraction speed largely dependent on their motor units and myosin proteins that can change in response to genetic and environmental factors. Human masticatory muscles express unique myosin isoforms, including a combination of thick fibers, expressing myosin light chains (MyLC) and myosin class I and II heavy chains (MyHC) -IIA, -IIX, α-cardiac, embryonic and neonatal and thin fibers, respectively. In this review, we discuss the current knowledge regarding the importance of fiber-type diversity in masticatory muscles versus supra- and infrahyoid muscles, and versus limb and trunk muscles. We also highlight new information regarding the adaptive response and specific genetic variations of muscle fibers on the functional significance of the masticatory muscles, which influences craniofacial characteristics, malocclusions, or asymmetry. These findings may offer future possibilities for the prevention of craniofacial growth disturbances.

Sarcoglycan immunoreactivity is lacking in infantile hypertrophic pyloric stenosis. A confocal laser scanning microscopic study.

OBJECTIVES: The Dystrophin-Glycoprotein Complex (DGC) is a large multisubunit complex that plays a crucial role in maintaining the structural integrity and physiology of muscle fibers. Dystrophin has been reported to be absent in the pyloric muscle of infantile hypertrophic pyloric stenosis (IHPS) patients. The present study was designed to investigate the other two patterns of DGC (dystroglycan and sarcoglycan complexes) in normal pyloric muscle and their possible modifications in IHPS patients. METHODS: Ten pyloric muscle biopsies were obtained from babies operated for IHPS and five control pylorus biopsy taken at autopsy from cases without gastrointestinal disease. The DGC sub-complexes (beta-dystroglican and beta, delta- sarcoglycans) were localized immunohistochemically using specific monoclonal antibodies. The results were evaluated using a confocal laser scanning microscope. RESULTS: Positive immunolocalization of the two DGC sub complexes was demonstrated in the smooth muscle cells (SMCs) of the pyloric region of control patients. Similarly, a positive immune expression of beta-dystroglican was observed in the pyloric SMCs of IHPS patients. On the other hand a negative immunoreaction for sarcoglycans was recorded within the full thickness of the pyloric SMCs of these patients. CONCLUSIONS: The absence of sarcoglycans within the hypertrophied pyloric muscle may be a predisposing factor in the pathogenesis of IHPS since it could alter the normal physiology of SMCs through the modifications of structural integrity of sarcolemma and signaling between the extracellular and intracellular compartment.

Integrins, muscle agrin and sarcoglycans during muscular inactivity conditions: an immunohistochemical study.

Sarcoglycans are transmembrane proteins that seem to be functionally and pathologically as important as dystrophin. Sarcoglycans cluster together to form a complex, which is localized in the cell membrane of skeletal, cardiac, and smooth muscle. It has been proposed that the dystrophin-glycoprotein complex (DGC) links the actin cytoskeleton with the extracellular matrix and the proper maintenance of this connection is thought to be crucial to the mechanical stability of the sarcolemma. The integrins are a family of heterodimeric cell surface receptors which play a crucial role in cell adhesion including cell-matrix and intracellular interactions and therefore are involved in various biological phenomena, including cell migration, and differentiation tissue repair. Sarcoglycans and integrins play a mechanical and signaling role stabilizing the systems during cycles of contraction and relaxation. Several studies suggested the possibility that integrins might play a role in muscle agrin signalling. On these basis, we performed an immunohistochemical analyzing sarcoglycans, integrins and agrin, on human skeletal muscle affected by sensitive-motor polyneuropathy, in order to better define the correlation between these proteins and neurogenic atrophy due to peripheral neuropathy. Our results showed the existence of a cascade mechanism which provoke a loss of regulatory effects of muscle activity on costameres, due to loss of muscle and neural agrin. This cascade mechanism could determine a quantitative modification of transmembrane receptors and loss of alpha7B could be replaced and reinforced by enhanced expression of the alpha7A integrin to restore muscle fiber viability. Second, it is possible that the reduced cycles of contraction and relaxation of muscle fibers, during muscular atrophy, provoke a loss of mechanical stresses transmitted over cell surface receptors that physically couple the cytoskeleton to extracellular matrix. Consequently, these mechanical changes could determine modifications of chemical signals through variations of pathway structural integrins, and alpha7A could replace alpha7B.

Sarcoglycans in human skeletal muscle and human cardiac muscle: a confocal laser scanning microscope study.

Sarcoglycans are a subcomplex of transmembrane proteins which are part of the dystrophin-glycoprotein complex. They are expressed in the skeletal, cardiac and smooth muscle. Although numerous studies have been conducted on the sarcoglycan subcomplex in skeletal and cardiac muscle, the manner of the distribution and localization of these proteins along the nonjunctional sarcolemma is not clear. We therefore carried out an indirect immunofluorescence study on surgical biopsies of normal human skeletal muscle and of healthy human atrial myocardium biopsies of patients affected by valvulopathy. Our results indicate that, in skeletal muscle, sarcoglycans have a costameric distribution and all colocalize with each other. Only in a few cases did the alpha-sarcoglycan not colocalize with other sarcoglycans. In addition, these glycoproteins can be localized in different fibers either in the regions of the sarcolemma over band I or band A. In cardiac muscle, our results show a costameric distribution of all proteins examined and, unlike in skeletal muscle, they show a constant colocalization of all sarcoglycans with each other, along with a consistent localization of these proteins in the region of the sarcolemma over band I. In our opinion, this situation seems to confirm the hypothesis of a correlation between the region of the sarcolemma occupied by costameric proteins and the metabolic type, fast or slow, of the muscular fibers. These data, besides opening a new line of research in understanding interactions between the sarcoglycans and other transmembrane proteins, could also be extended to skeletal and cardiac muscles affected by neuromuscular and cardiovascular pathologies to understand possible structural alterations.

Histochemical and biochemical analysis of phospholipase C isoforms in normal human gastric mucosa cells.

The expression and activity of PIP2-specific phospholipase C (PLC) in healthy human gastric mucosa cells were investigated by means of Western blotting, immunohistochemistry and in vitro activity assays. The results provide direct evidence for an almost exclusive expression of the PLC beta family and at the same time supply a cellular cartography of each represented isoform of this family. In this context, the putative roles of each isoform in the signaling events regulating the gastric mucosa metabolic machinery are discussed. These data provide a unique map of the specific expression and cellular distribution of the most represented PLC isoforms in healthy human gastric mucosa cells, which may constitute a reference point in future studies aimed at highlighting possible cytochemical and biochemical hallmarks of metaplastic or malignant transformation.

Nitric oxide production is increased in the spermatic veins of adolescents with left idiophatic varicocele.

BACKGROUND/PURPOSE: The existence of an excessive release of nitric oxide (NO) within dilated spermatic veins has been recorded in adults with varicocele suggesting a high oxidative stress. The authors investigated whether NO overproduction is already present in the dilated veins of adolescent varicocele and which enzymatic isoforms in the spermatic vein could be expressed. METHODS: The study group consisted of 10 adolescent patients affected by left idiophatic varicocele of grade II and III. The increase in NO production was established by determination of serum concentration of L-hydroxyarginine (L-NHA) and Nitrite/nitrate (NOx). Both endothelial and inducible NOsynthase (NOS) were investigated by Western blot analysis and by immunohistochemical localization using specific monoclonal fluorescein conjugated antibodies. RESULTS: Serum L-NHA levels were significantly greater in the spermatic veins when compared with the peripheral veins 176.8 +/- 32.3 micromol/L versus 3.38 +/- 0.64 micromol/L (P =.0004 Similarly, NOx levels were increased, respectively, 68.2 +/- 16.7 nmol/mL versus 12.9 +/- 2.65 nmol/mL (P =.029). Endothelial NOS was localized in the spermatic vein of varicocele patients, but not overexpressed; the inducible isoform was not detected. CONCLUSIONS: Adolescents with varicocele already present an increase in NO within dilated veins. The dilated spermatic vein is not the major source for the increase in NO level. These results could have an implication in the natural history of adolescent varicocele and in programming the ideal time for surgical treatment.

Nitric oxide synthase patterns in normal and varicocele testis in adolescents.

OBJECTIVE: To determine if the testes of normal adolescents can produce nitric oxide (NO), by assessing NO synthase (NOS) activity, and whether this activity changes in adolescents with left idiopathic varicocele. PATIENTS AND METHODS: After obtaining informed consent, testicular biopsies were obtained from eight adolescents (mean age 16.4 years; controls) who underwent surgery for inguinal hernia or hydrocele, and from 20 adolescents (mean age 16.2 years) operated for left idiopathic varicocele. Inducible and endothelial NOS (iNOS and eNOS) isoforms were investigated in the biopsy specimens by immunohistochemical localization and Western blot analysis using specific fluorescein-conjugated antibodies. RESULTS: Both normal and pathological samples expressed eNOS at the level of vessels and Leydig cells. The iNOS was expressed in Leydig cells of normal testes and over-expressed in Leydig cells of varicocele testes. CONCLUSION: Leydig cells of adolescent testes constitutively express iNOS. Under pathological conditions, e.g. varicocele, iNOS is up-regulated and is a possible source of NO overproduction. These results could be useful in explaining the pathogenesis of both testis and sperm dysfunction in varicocele.

Phosphatidylinositol-3-kinase activation and atypical protein kinase C zeta phosphorylation characterize the DMSO signalling in erythroleukemia cells.

Here we provide evidence for a role of phosphatidylinositol-3-kinase (PI-3-kinase) and for its product phosphatidylinositol-3,4, 5-triphosphate (PI3,4,5P3) in the occurrence of the metabolic differentiation state induced by DMSO in murine Friend erythroleukemia cells. Of note, the activation of PI-3-kinase correlated with the modulation of the activation of another enzyme, the atypical protein kinase C zeta (aPKC zeta). In particular, the expression of PI-3-kinase was substantially unaffected by DMSO treatment while its phosphorylation and the production of PI3,4,5P3 was strongly increased within 24 h of DMSO. Such a result was paralleled by an evident phosphorylation of a PKC zeta. Treatment of the cells with the two unrelated PI-3-kinase inhibitors wortmannin and LY 294002 impaired the recovery of the number of differentiated cells, therefore indicating that PI-3-kinase might be involved in the induction of erythroid differentiation, possibly engaging a protein kinase C zeta as downstream effector.

Differential effects of stromal derived factor-1 alpha (SDF-1 alpha) on early and late stages of human megakaryocytic development.

Stromal derived factor-1 alpha (SDF-1 alpha), the high-affinity ligand of CXC-chemokine receptor 4 (CXCR4), was added to human CD34(+) hematopoietic progenitor cells that can be induced to differentiate along the monocytic or megakaryocytic lineages. In control liquid cell cultures supplemented with two different cytokine cocktails: stem cell factor (SCF), interleukin-3 (IL-3), macrophage-colony stimulating factor (M-CSF), and 10% fetal calf serum (FCS), or, SCF and thrombopoietin (TPO), the expression of surface CXCR4 progressively increased in both the CD14(+) monocytic and CD41(+) megakaryocytic lineages. While SDF-1 alpha caused only modest effects on cells of the monocytic lineage, it induced profound down-regulation of CXCR4 in megakaryocytic cells at all stages of differentiation. Moreover, while SDF-1 alpha initially up-regulated the early megakaryocytic antigen CD41, at later time points (days 12-16) it induced down-regulation of the late megakaryocytic antigen CD42b. Consistently, at day 16, the number of mature megakaryocytes was significantly decreased in cultures supplemented with SDF-1 alpha. These findings indicate that, besides its primary role in regulating the retention of precursor cells in hematopoietic tissues, the SDF-1 alpha/CXCR4 system participates in the regulation of megakaryocytic development by stimulating the formation of immature megakaryoblasts and inhibiting the formation of mature megakaryocytes.