Developmental potentials of hematopoietic and neural stem cells following injection into pre-implantation blastocysts.

Pluripotent embryonic stem (ES) cells are able to differentiate in vivo into all cell types of the fetal and adult organism and in vitro they can differentiate into a variety of cell types. In contrast, multipotent somatic stem cells (SSCs) isolated from fetal and adult tissues differentiate into mature effector cells of their tissue. However, recent studies imply that SSCs can also generate cell types of heterologous tissues indicating unexpected broad differentiation potentials. In order to examine and compare the developmental potentials of SSCs, we exposed hematopoietic stem cells (HSCs) and neural stem cells (NSCs) to an environment that is permissive for the development of all cell types of the embryo, namely the mouse preimplantation blastocyst. Using this approach we were able to detect progeny of HSCs and NSCs frequently in developing chimeric animals. Analysis of 18 different adult tissues revealed minor preferences of HSCs for hematopoietic tissues, while progeny of NSCs were mostly detected in neural tissues. Furthermore we observe that human cord blood-derived CD34+ and CD34+/CD38- HSCs also engraft murine embryos and that human donor contribution persists into adulthood. Our studies show the existence of tissue specific engraftment preferences of HSCs and NSCs and that both stem cell types are non-ES cell-like.

Prevention of distal embolization during coronary angioplasty in saphenous vein grafts and native vessels using porous filter protection.

BACKGROUND: Although distal embolization and the "no-reflow" phenomenon are well described in saphenous vein graft (SVG) interventions, the frequency, magnitude, and characterization of embolized debris have not been evaluated in routine coronary interventions. A unique embolus protection device described herein provides a means of containing and retrieving plaque material dislodged during percutaneous coronary interventions. This report details the first clinical experience of the effectiveness and safety of an emboli protection system in 11 SVG lesions and 15 native coronary artery lesions. METHODS AND RESULTS: The AngioGuard Emboli Capture Guidewire (Cordis) consists of a PTCA wire with an expandable filter at the distal tip. The porous membrane permits normal distal blood flow, while trapping potential emboli by filtration. After crossing the lesion, the filter is expanded, and routine angioplasty is performed over the same wire. Emboli retrieval is achieved by collapsing the filter and retracting the emboli capture wire (ECW). In 26 patients, standard angioplasty was performed over the ECW; 20 of these 26 patients received a stent. Collected debris was sent for histopathological analysis. Plaque debris was retrieved after native coronary and SVG interventions in all cases. The ECW was positioned and retrieved without complications. No major adverse events occurred. Myocardial infarctions and no-reflow were not observed. CONCLUSIONS: The embolization of plaque fragments frequently occurs during coronary and SVG intervention. Distal embolization leading to microvascular obstruction and no-reflow could be successfully minimized by using the ECW.

Immunotherapy with nondepleting anti-CD4 monoclonal antibodies but not CD28 antagonists protects islet graft in spontaneously diabetic nod mice from autoimmune destruction and allogeneic and xenogeneic graft rejection.

BACKGROUND: T-cell activation and the subsequent induction of effector functions require not only the recognition of antigen peptides bound to MHC molecules by T-cell receptor (TCR) for antigen but also a costimulatory signal provided by antigen presenting cells. CD4 T-cell activation and function require the CD4 molecule as a coreceptor of TCR. The CD28/B7 pathway is a major costimulatory signal for T-cell activation and differentiation. METHODS: The effect of targeting CD4 by nondepleting anti-CD4 monoclonal antibodies (mAbs) versus blocking CD28/B7 by CTLA4Ig, anti-CD80 mAbs, and anti-CD86 mAbs on the prevention of recurrence of autoimmune diabetes after MHC-matched nonobese diabetes-resistant (NOR) islet transplantation in nonobese diabetic (NOD) mice were compared. Whether nondepleting anti-CD4 mAbs prolong allogeneic islet graft survival and xenogeneic pig islet graft survival in diabetic NOD mice were studied. Furthermore, the effect of nondepleting anti-CD4 mAbs combined with CTLA4Ig on allogeneic islet graft survival in NOD mice was investigated. RESULTS: Recurrence of autoimmune diabetes can be prevented by nondepleting anti-CD4 mAbs. Blocking the CD28/B7 costimulatory pathway by CTLA4Ig or by anti-CD80 mAbs and anti-CD86 mAbs cannot prevent recurrence of autoimmune diabetes after islet transplantation. Short-term treatment with nondepleting anti-CD4 mAbs significantly prolongs allogeneic islet graft survival and xenogeneic pig islet graft survival in diabetic NOD mice. But nondepleting anti-CD4 mAbs combined with CTLA4Ig decreased allogeneic islet graft survival. CONCLUSIONS: Nondepleting anti-CD4 mAbs but not CD28 antagonists protect islet grafts in diabetic NOD mice from autoimmune destruction and allogeneic and xenogeneic graft rejection. The efficacy of nondepleting anti-CD4 mAbs is compromised when it combines with CTLA4Ig.

GM1-gangliosidosis in Alaskan huskies: clinical and pathologic findings.

Three Alaskan Huskies, two females and one male, were diagnosed with GM1-gangliosidosis. Clinically, diseased animals exhibited proportional dwarfism and developed progressive neurologic impairment with signs of cerebellar dysfunction at the age of 5-7 months. Skeletal lesions characterized by retarded enchondral ossification of vertebral epiphyses were revealed by radiographs of the male dog at 5.5 months of age. Histologic examination of the central nervous system (CNS) revealed that most neurons were enlarged with a foamy to granular cytoplasm due to tightly packed vacuoles that displaced the Nissl substance. Vacuoles in paraffin-embedded sections stained positively with Luxol fast blue and Grocott's method, and in frozen sections vacuoles were periodic acid-Schiff positive. Foamy vacuolation also occurred within neurons of the autonomic ganglia. Extracerebral cells such as macrophages and peripheral lymphocytes also displayed foamy cytoplasm and vacuolation. In the CNS of diseased animals, a mild demyelination and axonal degeneration was accompanied by a significant astrogliosis (P < 0.05) in the gray matter as compared with age- and sex-matched control dogs. There was also a significant loss (P < 0.05) of oligodendrocytes in the gray and white matter of affected animals as compared with controls. Ultrastructurally, the neuronal storage material consisted of numerous circular to concentric whorls of lamellated membranes or stacks of membranes in parallel arrays. GM1-gangliosidosis in Alaskan Huskies resembles beta-galactosidase deficiency in other canine breeds, and these CNS disorders may be a consequence of neuronal storage and disturbed myelin processing.

Expression pattern of Oct-4 in preimplantation embryos of different species.

POU transcription factors are involved in transcriptional regulation during early embryonic development and cell differentiation. Oct-4, a member of this family, has been shown to be under strict regulation during murine development. The expression of Oct-4 correlates with the undifferentiated cell phenotype of the mouse preimplantation embryo. In this study, expression of a gene construct consisting of selected parts of the region upstream from the murine Oct-4 gene as promoter/enhancer, enhanced green fluorescent protein (EGFP) as reporter and the five exons of the murine Oct-4 gene (GOF18-delta PE EGFP) was evaluated in murine, porcine, and bovine preimplantation embryos. For comparison, expression of the endogenous Oct-4 gene was also analyzed in all three species by immunocytochemistry. The transgene construct was microinjected into zygotes cultured in vitro to various developmental stages. The EGFP fluorescence was visualized in developing embryos by excitation with blue light at different days following microinjection and showed similar expression patterns in all three species. Most embryos displayed a mosaic pattern of transgene expression. The EGFP fluorescence was not restricted to the inner cell mass (ICM) but was also seen in trophoblastic cells. An affinity-purified polyclonal antibody specific to Oct-4 was used for immunocytochemical analysis of in vivo- and in vitro-derived bovine and porcine blastocysts and also of in vivo-derived murine blastocysts. In the in vivo-derived murine embryos, Oct-4 protein was detectable in the ICM but not the trophectoderm, whereas in porcine and bovine blastocysts, derived in vivo or in vitro, Oct-4 protein was detected in both the ICM and the trophectoderm. Thus, in the two large animal species, Oct-4 expression from the endogenous gene was clearly not restricted to the pluripotent cells of the early embryo. These results show that Oct-4 regulation differs between these species and that the presence of Oct-4 protein may not be sufficient for selection of undifferentiated cell lines in domestic animals.

Posttransplant nonfunction of canine islets in PVG rats deficient in complement component C6.

BACKGROUND: Discordant islet xenografts are immediately nonfunctional in nonimmunosuppressed recipients other than the mouse, a process called primary nonfunction. Although at present it is unknown whether complement is involved, complement might participate in the induction of primary nonfunction through a number of mechanisms. We investigated the potential role of the membrane attack complex of complement in primary nonfunction of transplanted xenoislets. METHODS: Canine islets were transplanted into both nonimmunosuppressed and immunosuppressed normocomplementemic and C6-deficient (C6D) PVG rats. Cyclosporine, rapamycin, deoxyspergualin, and mycophenolate mofetil were used for immunosuppression from day -3 to cessation of islet cell function. Serum glucose was measured at 6 hr after transplant and daily thereafter. Xenograft tissue sections were obtained at various times after transplant and stained for inflammatory cells and insulin. RESULTS: Canine islets grafted in nonimmunosuppressed C6D rats and normocomplementemic rats underwent primary nonfunction in all animals. The incidence of primary nonfunction in animals receiving a four-drug immunosuppressive regimen was 33% in the normocomplementemic rats but only 10% in the C6D rats. The mean functional islet survival time was 1.57+/-0.33 days in the normocomplementemic group and 2.70+/-0.67 days in the C6D group (P=0.38). The islet xenografts showed little difference in degree and composition of cell infiltration between normocomplementemic and C6D rats. CONCLUSION: The membrane attack complex does not appear to play a major role in primary nonfunction of canine islet xenografts in nonimmunosuppressed PVG rats. However, there was a lower incidence of primary nonfunction and a longer posttransplant survival time in immunosuppressed C6D rats, suggesting the membrane attack complex may play a minor role in recipients that are heavily immunosuppressed.

Multiple peripheral nerve sheath tumors in the small intestine of a horse.

Multiple neurofibromas, schwannomas, and hyperplastic enteric plexuses were observed in the distal jejunum and ileum of a 6-year-old pinto gelding. The animal was presented because of an acute episode of colic. Three meters of distal small intestine, partially incarcerated in the epiploic foramen, were surgically removed. Numerous tumor nodules up to 10 mm in diameter were found adjacent to a Meckel's diverticulum, predominantly located in the subserosa of a hypertrophic segment. Histologically, tumors were well demarcated and composed of interlacing fascicles formed by spindloid cells. Adjacent enteric plexuses were hyperplastic. Immunohistochemically, all tumors were positive for vimentin and S-100. Desmin immunoreactivity was only observed in larger tumors (> 500 microns). Glial fibrillary acid protein was demonstrated nearly exclusively in smaller ones. Immunostaining for neurofilament was restricted to entrapped ganglion cells. Based on conventional light microscopic examination and immunohistochemical evaluation, the lesion was diagnosed as multiple benign peripheral nerve sheath tumors in the small intestine.

Equine adenocarcinomas of the large intestine with osseous metaplasia.

Large intestinal adenocarcinoma with osseous metaplasia was diagnosed in two horses, a 15-year-old standard bred gelding and a 9-year-old Haflinger mare. Clinically, both animals had displayed weight loss and anaemia. A presumptive diagnosis of abdominal neoplasia was made and the horses were humanely killed. At necropsy, the gelding and the mare were found to have ulcerated tumours growing into the lumen of the caecum and colon, respectively. In the mare, the mass extended through the mesocolon and was evident in the left dorsal and ventral colon. Histopathologically, the tumours consisted of well-differentiated cords of single-layered columnar to cuboidal epithelial cells. Mitotic figures were very uncommon. In both lesions, well-formed bony spicules and osteoid were present in the fibrovascular stroma. The tumours were well-demarcated from surrounding mucosal tissue but had invaded the intestinal wall. Metastases were not observed.

[Intraosseous epidermoid cysts of the toe phalanx in dogs]. / Intraossäre Epidermoidzysten des Zehenendgliedes beim Hund.

Out of 1057 amputated canine toes with tumours, tumour-like or other lesions, 17 cases were histopathologically diagnosed as epidermoid cysts of the terminal phalanx (EP). They were accompanied by osteolysis, reactive bone formation and chronic inflammation of phalanx 3. Extension of reactive changes to the phalangeal joint and phalanx 2 was less frequent. Various breeds were involved. There was no breed predisposition. The average age was 10.8 years; the male-female ratio 2.4:1. Nine EP were located on the forelimbs, 4 on the hindlimbs. In 4 cases location was unknown. Toes 4 and 5 were most frequently affected. A traumatic origin is suspected.