Cryopreserved porcine tendons preserve cell viability after thawing.

Controlled cryopreservation is an important method for storage of tissue grafts in skin banking, reproductive medicine and other domains. Although the availability of cryopreserved flexor tendons would be highly beneficial in reconstructive surgery, especially in complex reconstructions for which grafting material is limited, only a few studies have dealt with transplanted tendons. We achieved successful cryopreservation of porcine flexor tendons in 2 cryoprotective media: dimethyl sulfoxide and glycerol. Their viability was shown using a quantitative colorimetric MTT (3-[4,5-dimethyl-2-thiazolyl]-2,5-diphenyl-2H-tetrazolium bromide) assay. For comparison of native and cryopreserved tendons (n = 7 samples each), the adopted viability index was the ratio of MTT-dependent optical density and tendon weight. The viability index of native samples did not change significantly after cryopreservation and thawing. The proliferative capacity of tendon fibroblasts after thawing was shown in primary cell cultures. The described cryopreservation protocol and MTT assay may provide a basis for future autografting of human tendons.

[Nasal dermoid sinus cyst: accidental coincidence or syndrome association?]. / Nasendermoide bei Peters-Plus-Syndrom: Zufall oder Syndromexpression?

BACKGROUND: Peters anomaly is a rare congenital glaucoma disease. The Peters' plus syndrome is characterized by distinct malformations. As some of the common craniofacial malformations like cleft lip and palate are frequent in Peters' plus syndrome, no nasal dermoid sinus cysts has been reported so far. Nasal dermoid sinus cysts usually present in isolation, although associations to other anomalies or syndromes are possible. The occurrence of such an anomaly may be either accidental, or present a syndrome association. PATIENTS AND METHOD: One patient with an unilateral cleft lip and Peters' plus syndrome had undergone removal of nasal dermoid sinus cyst previously and was referred for management of recurrent disease. Complete surgical removal and plastic reconstruction was performed. RESULTS: Concerning the common (lateral) cleft lip nasal deformity with no midline nasal masses, there are reasons for the assumption that a coincidence of both anomalies might be accidental. Especially in Peters' plus syndrome no occurrence of nasal dermoids has thus far been documented. However, the embryological pathway of the frontonasal region differs from lip and palate development in time and location: So unique formation of both lesions seems inconsistent. Complete surgical removal and plastic reconstruction simultaneously or in a second step are recommended. CONCLUSION: As two cases of arhinia and Peters anomaly have been described in 1978, midline nasal masses might be a possible appearance of Peters' plus syndrome.

[Nasal dermoid and sinus cysts in patients with cleft malformations]. / Nasenfisteln und -zysten bei Patienten mit Lippen-Kiefer-Gaumen-Spalten.

BACKGROUND: Nasal dermoid sinus cysts are uncommon congenital lesions. They are usually isolated occurrences and are not associated with syndromes or additional malformations. The coincidence of both, cleft malformations and nasal dermoid sinus cysts, has seldom been reported. CASE REPORTS: Within the last 2 years two patients with reconstructed cleft lip and palate and additional nasal dermoid sinus cysts underwent surgical removal. One patient with bilateral complete cleft lip exhibited a fistula from the medial third of the nasal dorsum up to the glabella. Another patient with unilateral cleft lip and Peters' plus syndrome had undergone removal of a nasal dermoid sinus cyst 12 years ago and was referred for management of recurrent disease. DISCUSSION: Concerning the common cleft-dependent nose malformations with no midline nasal masses, there are reasons for the assumption that a coincidence of both anomalies might be accidental. Especially in Peters' plus syndrome no frequent occurrence of nasal dermoids has thus far been documented. However, the proximity and temporal closeness of an embryological pathway of the frontonasal region and lip development could also argue for a unique formation of both lesions. Complete surgical removal and plastic reconstruction simultaneously or in a second step are recommended.

Graves' disease and Hashimoto's thyroiditis in monozygotic twins: case study as well as transcriptomic and immunohistological analysis of thyroid tissues.

OBJECTIVE: To report on the rare simultaneous occurrence of Graves' disease (GD) and Hashimoto's thyroiditis (HT) in monozygotic twins. DESIGN: We compared the pattern of thyroid tissue-derived cDNAs to gain insight into previous and ongoing immune destruction and reconstruction processes using microarrays. The results were confirmed by immunohistology and real-time PCR. RESULTS: Destruction of thyroid tissue in HT reduced levels of thyrocyte-related cDNAs and cDNAs encoding extracellular matrix components, but increased levels of proteases involved in extracellular matrix degradation compared with GD. Lymphocytic infiltrates forming ectopic follicles replaced the thyroid tissue almost completely in HT. Thus, lymphocyte-related cDNA levels were higher in HT than in GD. The same was true for many chemokines and their receptors, which not only enable migration towards the thyroid but also maintain the lymphocytic infiltrate. HT also showed increased levels of cDNAs encoding molecules related to apoptosis than did GD. Surprisingly, the Th1- and Th2-specific cytokine profiles suggested for HT and GD respectively could not be confirmed. cDNAs encoding factors and receptors involved in angiogenesis were increased in GD compared with HT. CONCLUSIONS: Comparison of gene expression reflects the cellular differences between the two types of autoimmune thyroid disease in twins with identical genetic and similar environmental background.

Microvascular endothelial cells differ in their basal and tumour necrosis factor-alpha-regulated expression of adhesion molecules and cytokines.

We recently located a rare cytokeratin-positive (CK+) type of microvascular endothelial cell (MVEC) in the corpus luteum and aorta. Bovine corpus luteum MVEC are known to be involved in the cyclic accumulation of eosinophils and macrophages. Since leukocyte migration is specifically mediated by adhesion molecules and the release of cytokines, we compared the expression of these factors in basal and TNF-alpha-stimulated CK+ MVEC and in common cytokeratin-negative (CK-) MVEC in order to obtain an initial insight into the functional capacities of CK+ MVEC. CK- MVEC revealed significantly higher basal RANTES mRNA expression than CK+ MVEC, and TNF- alpha up-regulated RANTES mRNA in both types of MVEC. Only resting and stimulated CK- MVEC expressed granulocyte-macrophage colony-stimulating factor mRNA. Both MVEC types expressed monocyte colony-stimulating factor mRNA, but remained negative for eotaxin and interleukin (IL)-5 mRNA even after stimulation. Resting CK+ MVEC were positive for CD29, CD31, CD49a and CD49e, but expressed most of these antigens at a significantly lower density than did CK- MVEC. In contrast to CK- MVEC, CK+ MVEC failed to express CD49b or MHC class II. The activation of CK+ MVEC with TNF-alpha induced the expression of CD62P, but not of CD49b or MHC class II. In summary, phenotypically variable MVEC derived from the microvascular bed of one organ differ in their TNF-alpha-regulated expression of cytokine mRNA and adhesion molecules. Morphological heterogeneity is related to a particular specialisation of functional MVEC.

Retinoblastoma protein in microphthalmic mice.

A microphthalmic strain of mice was used to study immunoresponse of the retinoblastoma protein. Comparing wild-type, heterozygote and homozygote microphthalmic eyes, we found an increasing labelling of phosphorylated retinoblastoma protein (pRb) in the retinal pigment epithelium. Additionally, microphthalmic eyes expressed pRb in the neuroepithelium. Especially rosettes were strongly labelled.

Cloning of bovine RANTES mRNA and its expression and regulation in ovaries in the periovulatory period.

RANTES may be one of the chemoattractants involved in stimulating eosinophils and macrophages to migrate selectively into bovine dominant follicles and into developing corpora lutea. We sequenced a 736 bp fragment of the bovine RANTES mRNA encoding the complete protein and defined the ovarian source of RANTES mRNA. As demonstrated by competitive RT-PCR, follicle-derived macrophages showed a 100-1000 times higher RANTES mRNA level compared to unpurified granulosa cells or follicle-derived fibroblasts. By means of in situ hybridization, RANTES mRNA positive macrophages were located in the former thecal layer of the developing corpora lutea.

Heat shock proteins and heat shock protein-antibody complexes in placental tissues.

OBJECTIVE: The relationship between pregnancy outcome and expression of the heat shock proteins (hsps) or hsp-antibody complexes of 60kD (hsp60), 70kD (hsp70), and 90kD (hsp90) in placental tissue and circulating antibodies to hsps was evaluated. METHOD: Expression of hsp60, hsp70, and hsp90 in placentae from 12 women with preterm birth, eight with intrauterine growth restriction (IUGR), and 10 with term birth, as well as the presence of the corresponding antibodies, was investigated by a new carbocyanine double fluorescence technique. Results were compared with microbiological findings and circulating antibodies to hsps in sera. RESULTS: In each placental specimen examined, hsp60, hsp70, and hsp90 were identified. However, hsp70-antibody complexes were detected in only four of the preterm labor cases. Similarly, hsp60-antibody complexes were detected in only five preterm labor patients and in one patient with IUGR. None of the placentae contained hsp90-antibody complexes. In the preterm birth group, all patients with hsp60-antibody complexes were also positive for circulating antibodies to hsp60. The presence of hsp70-antibody complexes also correlated with hsp70 antibody in sera. CONCLUSIONS: Formation of hsp60- and hsp70-antibody complexes in the placenta may contribute to the induction of preterm birth. Women sensitized to these antibodies may be at increased risk for adverse pregnancy outcome.

Immunohistochemical detection of neuronal plexuses and nerve cells within the upper urinary tract of pigs.

OBJECTIVE: To investigate the distribution and topography of nervous structures within the renal pelvis and upper part of the ureter of pigs, and thus help to determine the origin, propagation and mechanisms of the modulation of pelvi-ureteric peristalsis. MATERIALS AND METHODS: Whole-mount preparations of renal pelves from adult pigs were stained using a universal immunostaining method with streptavidin-alkaline phosphatase. Anti-neuron-specific enolase antibody and anti-neurofilament antibody were used as neuronal markers. RESULTS: The patterns of neuronal structures differed between the renal calyces, renal pelvis and upper ureter. In the calyx, there was one single dense nerve plexus; this network contained relatively thin nerve fibres running both circularly and longitudinally. In the wall of the renal pelvis and upper ureter there were two neuronal plexuses, one submucosal and another within the muscular layer; these nerve fibres were mainly orientated longitudinally. Some single nerve cells were also found at the pelvicalyceal border. CONCLUSIONS: These findings suggest a potent nervous system within the upper urinary tract of pigs that connects the renal calyces with the renal pelvis, pelvi-ureteric junction and ureter. The presence of these dense neuronal networks and nerve cells within the wall of the renal pelvis and ureter suggests that propagation, coordination and modulation of pelvi-ureteric peristalsis in pigs may arise through intrinsic nervous stimulation.

Cytoarchitecture of the tectum opticum in the Japanese quail.

The cytoarchitecture of the optic tectum of the Japanese quail, Coturnix coturnix japonica, was studied using the Golgi-Kopsch method, parvalbumin, calbindin and GABA immunohistochemistry and nicotinamide adenine dinucleotide phosphate-diaphorase histochemistry. Our results reveal a large number of different types of interneurons in the quail tectum opticum, only part of which are described in the chick or pigeon. Application of parvalbumin and calbindin immunohistochemistry and nicotinamide adenine dinucleotide phosphate-diaphorase histochemistry reveals the following lamination pattern: The stratum opticum, stratum griseum centrale and stratum album centrale remain unstained, while the laminae of the stratum griseum et fibrosum superficiale exhibit a roughly complementary staining pattern of calbindin (laminae c, d, e, f, g, i) and parvalbumin (laminae a, h, i). Nicotinamide adenine dinucleotide phosphate-diaphorase histochemistry yields a dense band in lamina i. The Golgi material reveals the following cell types in the stratum griseum et fibrosum superficiale: marginal cells in the stratum opticum and in lamina h and i, horizontal cells in laminae a and c, large and small radial cells in laminae b, d, h and i, multiform cells in lamina b, bitufted cells in lamina d and e, large pear-shaped cells in lamina g, wide-field cells in lamina j, and stellate cells in lamina j and in the stratum griseum centrale. We consider horizontal cells, bitufted cells, multiform cells and small radial cells to be GABAergic interneurons of the stratum griseum et fibrosum superficiale which seem to be more numerous than in the pigeon tectum opticum. Golgi impregnation and injection of Phaseolus vulgaris leucoagglutinin into the pretectal nucleus lentiformis yielded regularly distributed clusters of telodendra of pretectal axons in lamina d of the stratum griseum et fibrosum superficiale, which are identical in shape and position with axon plexus revealed by Golgi staining.

Aberrant morphology of Müller glial cells in retinas of a microphthalmic mouse strain.

A microphthalmic mouse strain was used to study retinal glial cells during postnatal development of the retinal malformations. Glia was demonstrated with immunohistology using antibodies against vimentin or glial fibrillary acidic protein. To identify proliferating cells the bromodeoxyuridine technique was applied. Our results support the hypothesis that precursors of Müller cells may be involved in early stages of retinal malformations.