Male infertility can result from impaired sperm motility caused by multiple morphological abnormalities of the flagella (MMAF). Distinct projections encircling the central microtubules of the spermatozoal axoneme play pivotal roles in flagellar bending and spermatozoal movement. Mammalian sperm-associated antigen 17 ( SPAG17 ) encodes a conserved axonemal protein of cilia and flagella, forming part of the C1a projection of the central apparatus, with functions related to ciliary/flagellar motility, skeletal growth, and male fertility. This study investigated two novel homozygous SPAG17 mutations (M1: NM_206996.2, c.829+1G>T, p.Asp212_Glu276del; and M2: c.2120del, p.Leu707*) identified in four infertile patients from two consanguineous Pakistani families. These patients displayed the MMAF phenotype confirmed by Papanicolaou staining and scanning electron microscopy assays of spermatozoa. Quantitative real-time polymerase chain reaction (PCR) of patients' spermatozoa also revealed a significant decrease in SPAG17 mRNA expression, and immunofluorescence staining showed the absence of SPAG17 protein signals along the flagella. However, no apparent ciliary-related symptoms or skeletal malformations were observed in the chest X-rays of any of the patients. Transmission electron microscopy of axoneme cross-sections from the patients showed incomplete C1a projection and a higher frequency of missing microtubule doublets 1 and 9 compared with those from fertile controls. Immunofluorescence staining and Western blot analyses of spermatogenesis-associated protein 17 (SPATA17), a component of the C1a projection, and sperm-associated antigen 6 (SPAG6), a marker of the spring layer, revealed disrupted expression of both proteins in the patients' spermatozoa. Altogether, these findings demonstrated that SPAG17 maintains the integrity of spermatozoal flagellar axoneme, expanding the phenotypic spectrum of SPAG17 mutations in humans.
Humans ; Male ; Infertility, Male/pathology* ; Sperm Tail/ultrastructure* ; Homozygote ; Microtubule-Associated Proteins/genetics* ; Axoneme/genetics* ; Spermatozoa/ultrastructure* ; Adult ; Mutation ; Sperm Motility/genetics* ; Pedigree ; Microtubules ; Microtubule Proteins/genetics*

This study was conducted to evaluate the microtubule distribution following control of nuclear remodeling by treatment of bovine somatic cell nuclear transfer (SCNT) embryos with caffeine or roscovitine. Bovine somatic cells were fused to enucleated oocytes treated with either 5 mM caffeine or 150 micrometer roscovitine to control the type of nuclear remodeling. The proportion of embryos that underwent premature chromosome condensation (PCC) was increased by caffeine treatment but was reduced by roscovitine treatment (p < 0.05). The microtubule organization was examined by immunostaining beta- and gamma-tubulins at 15 min, 3 h, and 20 h of fusion using laser scanning confocal microscopy. The gamma-tubulin foci inherited from the donor centrosome were observed in most of the SCNT embryos at 15 min of fusion (91.3%) and most of them did not disappear until 3 h after fusion, regardless of treatment (82.9-87.2%). A significantly high proportion of embryos showing an abnormal chromosome or microtubule distribution was observed in the roscovitine-treated group (40.0%, p < 0.05) compared to the caffeine-treated group (22.1%). In conclusion, PCC is a favorable condition for the normal organization of microtubules, and inhibition of PCC can cause abnormal mitotic division of bovine SCNT embryos by causing microtubule dysfunction.
Animals ; Caffeine/pharmacology ; Cattle/embryology/*physiology ; Cell Nucleus/drug effects/*physiology/ultrastructure ; Female ; Fertilization in Vitro/veterinary ; Male ; Microscopy, Confocal/veterinary ; Microtubules/drug effects/*physiology/ultrastructure ; Nuclear Transfer Techniques/veterinary ; Oocytes/*physiology ; Pregnancy ; Purines/pharmacology

OBJECTIVETo investigate the changes in the morphology and cytoskeleton (CSL) content of the CSL in the colonic smooth muscle cells (SMCs) of rats in early postscald stage, so as to elucidate the mechanism of dysfunction of gastrointestinal motility.

METHODSSeventy Wistar rats were randomly divided into normal control (n = 10, without scald) , and scald ( n = 60, with 10 cm x 7 cm wound inflicted on the back) groups. The colonic smooth muscle tissue of 10 normal rats and scalded rats were harvested at 1, 3, 6, 12, 24 and 48 postscald hours( PSH) and divided into two parts: one for histologic examination, and the other for the detection of CSL changes in the colonic smooth muscle tissue by flowcytometry method.

RESULTSThe electron microscope examination showed that the arrangement of cytoskeleton of SMC of the scalded rats during 1 to 3 PSH was disordered, and sparse, and the condensed area was uneven, with fragmentation. But the morphology and distribution of CSL gradually restored to normal state during 6 to 12 PSH, and it approached that of normal group at 48 PSH. The CSL content in the colonic smooth muscle tissue of scalded rats was obviously increased at 1 PSH (610+/-23) , decreased thereafter, evidently lower than that in control group at 3 PSH (92+/-17) , and then it started to increase at 12 PSH, exceeding the normal value at 24 PSH, and continued to rise until 48 PSH. There was significant difference in CSL content in the colonic smooth muscle tissue of the rats between scald and control group ( P < 0.05 or 0. 01).

CONCLUSIONChanges in the morphology and CSL content in the colonic smooth muscle tissue can be observed at early stage after a scald, which imply the kinetic balance between damage and repair in the body. In addition, changes in CSL content in the colonic smooth muscle tissue may be important factors in producing colonic dysfunction, damage of intestinal wall structure, and dynamic abnormalities of the colonic smooth muscle.

Actins ; metabolism ; Animals ; Burns ; metabolism ; pathology ; Colon ; cytology ; Cytoskeleton ; ultrastructure ; Disease Models, Animal ; Microtubules ; Myocytes, Smooth Muscle ; cytology ; metabolism ; Rats ; Rats, Wistar

We report a first Korean case of presumably dominantly inherited primary tubular aggregate myopathy in a 19-yr-old man, who presented with slowly progressive proximal muscle stiffness and weakness. In hematoxylin and eosin stain, it showed subsarcolemmal, or central pale basophilic granular vacuoles, which stained red with modified Gomori's trichrome and intensive blue with nicotinamide adenonine dinucleotide-tetrazolium reductase, respectively. Ultrastructurally, aggregates of 60 nm-sized hexagonal tubules were found in both type 1 and type 2 fibers. We briefly review the pathologic findings of the previously reported cases of tubular aggregate myopathy and discuss the possible pathogenesis of this disease. We briefly discuss the possible pathogenesis of sarcoplasmic reticulum and review the ultrastructural characteristics.
Adult ; Biopsy ; Frozen Sections ; Genes, Dominant ; Genes, Recessive ; Human ; Korea ; Male ; Microscopy, Electron ; Microtubules/ultrastructure ; Mitochondria, Muscle/ultrastructure ; Muscle, Skeletal/pathology* ; Myopathies, Structural, Congenital/diagnosis ; Myopathies, Structural, Congenital/genetics ; Myopathies, Structural, Congenital/pathology* ; Pedigree