Fine structural changes in the Achilles tendons of patients with long-term diabetes mellitus were investigated. All patients had clinical and electrophysiological evidence of diabetic neuropathy and had ulceration and/or Charcot neuroarthropathy. Several differences between tendons of diabetic (n = 12) and nondiabetic (n = 5) individuals were observed by electron microscopy. In diabetics, these differences included increased packing density of collagen fibrils, decreases in fibrillar diameter, and abnormal fibril morphology. In one diabetic patient, individual collagen fibrils were tightly apposed so that many areas of tendon appeared as a single mass of closely adhering fibrillae. In addition, foci in which collagen fibrils appeared twisted, curved, overlapping and otherwise highly disorganized were common in specimens from most patients (11 of 12). These morphologic abnormalities in the Achilles tendons of diabetics appear to reflect a poorly known process of structural reorganization that may be the result of nonenzymatic glycation expressed over many years. Such structural changes could contribute to the tightening of the Achilles tendor a phenomenon consistent with clinical observations of extreme shortening of the Achilles tendon-gastrocnemius-soleus complex common in advanced diabetic neuropaths. In patients with diabetic neuropathy, tendon shortening causes severe equinus that may precipitate serious ulceration, stress fractures, and Charcot collapse of the foot. However, in nondiabetics, the fine structure of the Achilles tendon appears normal, consistent with the finding that the ultrastructural changes result from diabetes rather than neuropathy.

Achilles Tendon↗

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Some vasovasostomized men are characterized by low levels of P34H, an epididymal sperm protein.

During epididymal transit, sperm surface proteins involved in the fertilization process can be added or modified. P34H, a human epididymal-sperm protein, is proposed to be involved in the interactions between spermatozoa and the zona pellucida. We have previously demonstrated that P34H is present in men of proven fertility and is absent in 50% of men presenting with idiopathic infertility. Spermatozoa with a low amount of P34H exhibit a dramatic reduction in their ability to interact with zona pellucida. Even if the surgical success of vasectomy reversal is high, fertility is not always reestablished, possibly due to epididymal damage caused by vasectomy. In this study, western blot analyses were performed to determine the level of P34H present on spermatozoa of men who underwent vasectomy reversal. Spermatozoa obtained from different semen samples from a given individual had similar P34H levels; however, samples from different men were highly variable. When quantified by densitometric scanning, P34H levels from vasovasostomized men varied between 1.5% and 149% compared with that from a fertile donor who represented 100%. Eighteen of 25 vasovasostomized men had a P34H level lower than 30% of the normal value, while the remaining 7 males were in the normal range. Furthermore, the population of vasovasostomized men with P34H levels lower than 30% was significantly different from the control group of 19 fertile men. The high variation of P34H levels observed in vasovasostomized men did not correlate with the spermiogram values (P > 0.05). An important factor in determining sperm P34H level appears to be the period of time elapsed between the vasectomy and vasovasostomy. In summary, our results show that the P34H level varied from one man to another and that low levels of the epididymal sperm protein is associated with vasectomy reversal.

Adult↗

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Determining a newborn's gestational age.

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Female↗

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Screening for and managing dysphagia after stroke.

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Algorithms↗

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[Social capital, health promotion and population health].

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Canada↗

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Cryopreservation alters the levels of the bull sperm surface protein P25b.

Fertility of frozen-thawed bull sperm is reduced by cryopreservation. Freezing-thawing procedures can result in as much as a sevenfold fertility decrease. Sperm mortality and loss of motility do not fully explain the reduced fertility of cryopreserved semen; they may be partially explained by the loss of sperm surface proteins, which are necessary for fertilization. We have previously identified P25b, a sperm surface protein, which is associated with the fertility index of bulls used for artificial insemination. Using Western blotting techniques, we have evaluated P25b levels before and after cryopreservation of bull spermatozoa in extenders based on either egg yolk or milk. Long storage periods (28 days) in liquid nitrogen results in a threefold decrease of P25b levels associated with cryopreserved versus fresh spermatozoa. Over a short storage period (3-7 days), a stable P25b level was observed on spermatozoa cryopreserved in extender containing either egg yolk or milk. A decrease in P25b levels associated with spermatozoa was observed after 5 days of storage in egg yolk extender, whereas a significant decrease was observed after 14 days of sperm storage in milk extender (P < .05). Therefore, the loss of P25b may be responsible, at least in part, for the decrease in fertility following the freezing-thawing procedure of bull semen. Moreover, the cryopreservation extender used may have different effects on the loss of sperm surface proteins after even brief storage periods in liquid nitrogen. Considering that a sperm protein similar to P25b exists in humans (P34H), these results may have significant clinical applications in which frozen semen is used.

Animals↗

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Epididymal epithelial cells cultured in vitro prolong the motility of bovine sperm.

It is well known that the epididymis is an excellent environment to maintain sperm viability. Therefore, we used different sections of bovine epididymis (caput, corpus, and cauda) to develop epithelial cell culture monolayers to identify factors that will increase sperm survival in the freezing-thawing process. Each epididymal section was dissected and treated with collagenase to obtain epithelial cell clusters. The cells were cultured in RPMI-1640 medium with 10% serum at 38.5 degrees C. A confluent monolayer was obtained after 5-7 days in culture and preliminary characterization using cytokeratin antibody indicated that the cell culture contained 85%-95% of epithelial cells. These cellular cultures were tested for their ability to maintain motility of epididymal and frozen-thawed spermatozoa. Washed spermatozoa were added to obtain a final dilution of 1 x 10(6) spermatozoa/mL. The motility of frozen-thawed spermatozoa was also recorded after incubation in conditioned media. Our results show that cocultures of spermatozoa and epididymal cell monolayers for 24 and 48 hours were beneficial for maintaining epididymal and frozen-thawed sperm motility (36.0% and 20.4%) compared with spermatozoa cultured with fibroblast cells or in the absence of a cell monolayer (0%; P < .01). The conditioned medium provides favorable conditions for sperm motility. Results with conditioned medium on maintenance of frozen-thawed sperm motility suggest that epididymal cells in vitro secrete beneficial factors that prolong the sperm survival.

Animals↗

PubMedDate not supplied

Addition of specific metabolites to bovine epididymal cell culture medium enhances survival and motility of cryopreserved sperm.

We have developed a cell culture system of bovine epididymal epithelium in which cryopreserved bovine sperm motility was efficiently maintained for many hours. The culture conditions to maintain viable epididymal cells are quite different from conditions normally used to incubate sperm cells. Thus, we have modified a previously described principal cell medium (PCM; Moore et al, 1992) using HEPES as a buffer and supplemented media with myo-inositol, pyruvate, lactate, glycerol, and carnitine to mimic epididymal intraluminal conditions. In the first experiments the effects of PCM and our epididymal cell medium (ECM) on sperm motility were compared in the absence of cells and evaluated by microscopic analysis under a phase contrast microscope or using the Hamilton Thorn Image Analyzer System. Our results showed that motility of cauda epididymal sperm was significantly higher in ECM than in PCM during a 48-hour incubation period when both media were supplemented with 10% fetal bovine serum (FBS). We then replaced FBS with bovine serum albumin (BSA) or no proteins at all to verify if ECM was able to enhance sperm survival. To test this aspect we used frozen-thawed sperm, which survived up to 48 hours when sperm cells were coincubated with epididymal cell monolayers. Hence, PCM, ECM, and different media containing each metabolite of ECM were supplemented with 0.5% BSA to assess motility of thawed sperm after an incubation period of 6 hours. A positive effect on sperm motility was observed in all fresh and unconditioned media containing 1 mM pyruvate. Motion parameters were more efficiently maintained in all conditioned media than in unconditioned media. Our results showed, however, that pyruvate was almost completely oxidized or consumed by epididymal cells during preincubation of culture media. We conclude that motility of frozen-thawed bovine spermatozoa can be improved using a culture medium or a medium conditioned by epididymal cell cultures without carnitine but containing mainly pyruvate, inositol, glycerol, and lactate.

Animals↗

PubMedDate not supplied

Detection of low numbers of poliovirus 1 in oysters: collaborative study.

A collaborative study was performed to evaluate a method for determining numbers of poliovirus 1 in oysters. Commonly available laboratory equipment and materials were used. Raw oysters in the shell were shipped to each investigator along with 12 tubes of unknown concentrations of virus. Six 100 g duplicate oyster samples were analyzed by 5 collaborating laboratories. Each analyst used a prescribed procedure for diluting the inocula. Two samples contained approximately 100 plaque-forming units (pfu)/100 g, 2 samples contained approximately 50 pfu/100 g, and the other 2 contained approximately 25 pfu/100 g. Recoveries varied from 35 to 55% at inocula levels of 30-100 pfu/100 g. The limit of detectability was hypothesized to be 14 pfu in a 100 g sample when the recovery was 35%. The method has been adopted official first action.

Animals↗