Reduction of donor site morbidity of the iliac crest free flap by preservation of the anterior superior iliac spine

To reduce donor site morbidity in the iliac crest free flap, we suggest leaving the anterior superior iliac spine in situ. The advantages are: less tension on the wound, less pain, faster rehabilitation, preservation of the ability to wear pants without braces, and a better cosmetic result through preservation of contour. Received: 1 December 1999 / Accepted: 1 February 2000     

Gpr177/mouse Wntless is essential for Wnt‐mediated craniofacial and brain development

We have previously demonstrated that Gpr177, the mouse orthologue of Drosophila Wls/Evi/Srt, is required for establishment of the anterior–posterior axis. The Gpr177 null phenotype is highly reminiscent to the loss of Wnt3, the earliest abnormality among all Wnt knockouts in mice. The expression of Gpr177 in various cell types and tissues lead us to hypothesize that reciprocal regulation of Wnt and Gpr177 is essential for the Wnt‐dependent developmental and pathogenic processes. Here, we create a new mouse strain permitting conditional inactivation of Gpr177. The loss of Gpr177 in the Wnt1‐expressing cells causes mid/hindbrain and craniofacial defects which are far more severe than the Wnt1 knockout, but resemble the double knockout of Wnt1 and Wnt3a as well as β‐catenin deletion in the Wnt1‐expressing cells. Our findings demonstrate the importance of Gpr177 in Wnt1‐mediated development of the mouse embryo, suggesting an overlapping function of Wnt family members in the Wnt1‐expressing cells. Developmental Dynamics 240:365–371, 2011. © 2011 Wiley‐Liss, Inc.     

Variations of the prominences of the bony palate and their relationship to complete dentures in Korean skulls.

The osteological and morphological variations of the prominences in the bony palate of 160 Korean skulls were studied. The frequency of the occurrence of the posterior palatine crest, located on the posterior border of the greater palatine foramen, was 13.8%. Palatal ridges were observed commonly in the skulls; however, the smooth type, which has no palatal ridges in the palate, was shown in 14.7% of cases, and palatal spines were observed in 33.8%. The prevalence of palatal tubercles was 11.6%, and all were found in the molar region. The palatine torus was found in 18.8% of cases and the most common type was along the median palatine suture from the incisive foramen to the posterior border of the palatine bone (63.3%). No significant differences between sexes or sides were found in the posterior palatine crest, palatal ridges, and palatal tubercle. However, the sex distribution of the palatine torus was significantly different (P < 0.05). These results would be helpful clinically in fabricating maxillary complete dentures for edentulous patients.     

The rostral dorsal cap and ventrolateral outgrowth of the rabbit inferior olive receive a GABAergic input from dorsal group Y and the ventral dentate nucleus

The dorsal cap and ventrolateral outgrowth of the inferior olive are involved in the control of eye movements. The caudal dorsal cap is predominantly involved in the horizontal optokinetic reflex; it receives most of its GABAergic input from the nucleus prepositus hypoglossi. In the present study, we determined the source of a major inhibitory input to the rostral dorsal cap and the ventrolateral outgrowth, which are the olivary subnuclei mainly involved in the “vertical” optokinetic reflexes. We studied these subnuclei in the rabbit with the use of retrograde tracing of horseradish peroxidase and anterograde tracing of wheat germ agglutinin-coupled horseradish peroxidase combined with postembedding immunocytochemistry. The ventral dentate nucleus of the cerebellum and dorsal group y project contralaterally to the rostral dorsal cap and ventrolateral outgrowth; this projection is entirely GABAergic. The terminals of this input form predominantly symmetric synapses with extraglomerular and intraglomerular dendrites; the remaining terminals are axosomatic. In addition, the dorsal cap and ventrolateral outgrowth contain significantly more crest synapses than any other olivary subnucleus. The terminals that form these crest synapses are derived from dorsal group y and/or the ventral dentate nucleus. None of the terminals in the dorsal cap or ventrolateral outgrowth was glycinergic.     

Rostro-caudal polarity in the avian somite related to paraxial segmentation

Segmental organization of the vertebrate body is one of the major patterns arising during embryonic development. Somites that play an important role in this process show intrinsic patterns of gene expression and differentiation. The somites become polarized in all three dimensions, rostrocaudal, mediolateral and dorsoventral, the quadrants giving rise to several tissue components. The timing of polarization was studied by means of antibodies against HNK-1, tenascin and neurofilament. Whole mounts and serial sections of quail and chick embryos show that somites are already polarized at the moment of their segregation from the segmental plate. The rostral hemisomite carries the HNK-1 epitope preferentially, while the caudal hemisomite stains more strongly for tenascin. HNK-1-stained areas in the segmental plate strongly relate to the notochordal sheath, suggesting that axial structures determine the fate of paraxial structures. Neural crest cells were only seen to colonize the rostral part of a somite after they had differentiated into HNK-1 positive cells. Their colonization pattern seems to be guided by the segmental organization of the somite. Moreover, this somite organization probably dictates the organization of both sensory and motor fibres converging towards the segmental dorsal root ganglia, justifying a shift in the connections between neural tube and somites. This segmental shift takes place over one quarter of a somite length in a rostral direction.     

Heparitinase treatment of rat embryos during cranial neurulation

Summary Heparan sulphate has been reported to be present in rat embryos. It is covalently linked to a core protein as heparan sulphate proteoglycan (HSPG). Heparitinase specifically degrades heparan sulphate, thus treatment of rat embryos with this enzyme in vitro should result in the perturbation of any tissue interactions which involve heparan sulphate proteoglycan. In this study heparitinase was either added to the culture medium or microinjected directly into the amniotic cavity.Heparitinase treatment resulted in abnormal development of the whole embryo, but the earliest effects were observed in the cranial region. Forebrain development was grossly abnormal: the neural folds remained widely open, with beak-like outgrowths rostrally. Optic sulci failed to develop. The midbrain and rostral hindbrain neural folds also remained widely open. In the trunk, where the pattern of neurulation is less complex than in the cranial region, rostral neural tube closure did occur although the morphology of the closed region was far from normal. These results suggest that heparan sulphate proteoglycan is essential for normal neurulation.Epithelial somite formation was perturbed, but neural crest cell emigration, otic pit formation and pharyngeal arch formation, all important morphogenetic events which occur during this period of development, were not inhibited by heparitinase treatment. Prolonged (44h) exposure to the enzyme resulted in the conversion of the embryonic structure to a much simpler form: mesenchymal cells (stellate or spindle-shaped) enclosed within a simple epithelial coating.     

小鼠颅神经嵴细胞的培养和特征

目的：在体外原代培养Balb/c小鼠胚胎的颅神经嵴细胞。为颅面部各种组织细胞的发育研究提供细胞来源。方法：采用胰酶消化法分离小鼠胚胎第8.5天的颅神经管，从小鼠颅神经管中游离出来的细胞即为颅神经嵴细胞，用免疫组织化学方法鉴定细胞的来源，并测定细胞的生长曲线。结果：成功地培养出小鼠的颅神经嵴细胞，其形态类似成纤维样细胞，免疫组化检测结果表明，神经特异性烯醇化酶（NSE）抗体染色结果阳性。细胞的群体倍增时间为43.65h。结论：原代培养的小鼠颅神经嵴细胞生长稳定，来源明确，是颅面部各种细胞的发育和分化研究中一种有用的工具。     

Human neural crest cells contribute to coat pigmentation in interspecies chimeras after in utero injection into mouse embryos

The neural crest (NC) represents multipotent cells that arise at the interphase between ectoderm and prospective epidermis of the neurulating embryo. The NC has major clinical relevance because it is involved in both inherited and acquired developmental abnormalities. The aim of this study was to establish an experimental platform that would allow for the integration of human NC cells (hNCCs) into the gastrulating mouse embryo. NCCs were derived from pluripotent mouse, rat, and human cells and microinjected into embryonic-day-8.5 embryos. To facilitate integration of the NCCs, we used recipient embryos that carried a c-Kit mutation (W^sh/W^sh), which leads to a loss of melanoblasts and thus eliminates competition from the endogenous host cells. The donor NCCs migrated along the dorsolateral migration routes in the recipient embryos. Postnatal mice derived from injected embryos displayed pigmented hair, demonstrating differentiation of the NCCs into functional melanocytes. Although the contribution of human cells to pigmentation in the host was lower than that of mouse or rat donor cells, our results indicate that hNCCs, injected in utero, can integrate into the embryo and form mature functional cells in the animal. This mouse–human chimeric platform allows for a new approach to study NC development and diseases.Genetically engineered mice have been highly informative in studying the developmental origin of many inherited diseases (1–3). However, mouse models often fail to reproduce the pathophysiology of human disorders due to interspecies divergence, such as metabolic differences between mouse and human (4), or differences in genetic background (5). To overcome some of the limitations of transgenic mouse models, transplantation of disease-relevant human cells into mice has been informative and is frequently used in cancer research. However, this approach is primarily restricted to the study of end-stage-disease cell types and provides only limited insight into tumor initiation and early progression of the disease under in vivo conditions, with the exception of the hematopoietic lineages, where human hematopoietic stem cells were found to successfully engraft into immune-deficient mice and provided a powerful approach for studying blood diseases (6).Somatic cell reprogramming provides patient-specific induced pluripotent stem cells (iPSCs) that carry all genetic alterations contributing to the disease pathophysiology and thus allows for generating the disease-relevant cell types in culture (7). However, many complex diseases involve progressive cellular or genetic alterations that occur before the manifestation of a clinical phenotype. Therefore, it is not clear whether a disease-relevant phenotype can be observed in short-term cultures of cells derived from patients with long-latency diseases, such as Parkinson''s or Alzheimer’s disease or cancers like melanoma. A major challenge is establishing model systems that, using human embryonic stem cells (hESCs) or hiPSCs, will allow for the investigation of human disease under appropriate in vivo conditions.Transplantation of hiPSCs or hiPSC-derived cells into mouse embryos would present an attractive solution to many of the aforementioned limitations. The main advantage of such an approach is that the transplanted cells would integrate into the embryo and participate in normal embryonic development, and consequently could be studied over the lifetime of the mouse. Currently, it is controversial whether the injection of hESCs/hiPSCs into preimplantation mouse blastocysts can generate even low-grade chimeric embryos (8–11). As an alternative approach, we explored whether multipotent somatic cells would be able to functionally integrate into postgastrulation mouse embryos and allow for the generation of mouse–human chimeras. We investigated the potential of human neural crest cells (hNCCs), derived from hESCs/hiPSCs, to integrate into the mouse embryo and contribute to the NC-associated melanocyte lineage. The NC, a multipotent cell population, arises at the boundary between the neuroepithelium and the prospective epidermis of the developing embryo. Trunk NCCs migrate over long distances, with the lateral migrating NCCs generating all of the melanocytic cells of the animal’s skin (12).NCC migration, development, and differentiation into various tissues have been studied in vivo by generating quail–chick NC chimeras. In this model, donor quail tissues were grafted into similar regions of developing chicken embryos (13). The experimental approach of our present study was based on the generation of mouse–mouse NC chimeras that had been created by injection of primary mouse NCCs into the amniotic cavity of embryonic-day (E) 8.5 embryos (14, 15). The donor mouse NCCs (mNCCs), having been placed outside of the embryo, enter into the neural tube, presumably through the still-open neural pores, and transverse the epidermis. The donor mNCCs used in this previous study were collected from pigmented C57BL/6 mice, whereas the host embryos were derived from BALB/c albino mice. Thus, contribution of the donor mNCCs to the host embryo could be determined by the presence of pigmentation in the coats of the injected mice. The injected primary mNCCs contributed to coat color formation in the head and hind limb regions only, but not in the midtrunk area, likely reflecting the entry point of the cells through the neural pores with the anterior–posterior movement of the cells being hindered by endogenous melanoblasts (15). Indeed, when embryos carrying the white-spotted c-Kit mutation (W^sh/W^sh), which lack melanoblasts, were used as a host, extensive coat color contribution revealing anterior–posterior cell migration was observed, presumably because the donor NCCs could spread into the empty niches (14).Here, we differentiated mouse, rat, and human ESCs or iPSCs into NCCs that were injected in utero into E8.5 albino wild-type and c-Kit–mutant W^sh/W^sh embryos. Both the mouse and human NCCs migrated laterally under the epidermis and ventrally into deeper regions of the embryo. Importantly, analysis of postnatal animals derived from mouse, rat, or human NCC-injected embryos displayed coat color pigmentation from the donor cells. Our results demonstrate that NCCs from different species can integrate into the developing mouse embryo, migrate through the dermis, and differentiate into functional pigment cells in postnatal mice. The generation of postnatal mouse–human chimeras carrying differentiated and functional human cells allows for a novel experimental system in which to study human diseases in an in vivo, developmentally relevant environment.