[Ultrastructural studies of hepatoblast junctions and liver hematopoiesis of the mouse embryo].

To clarify the morphological changes in hepatoblast connections during the development of fetal liver hematopoiesis, ICR mouse livers of 11 to 19 days of gestation were studied by means of three-dimensional reconstruction, immunohistochemistry, electron microscopy and freeze fracture replica method. Embryonic liver weight showed rapid increase until 19 days of gestation, and an initial steep increase, due to hematopoietic development, was observed at 13 to 15 days of gestation. Hepatoblast volume appeared to be constant until 13 days of gestation, and, thereafter, showed a gradual increase. An 11-day primitive hepatic cord contained a few immature hematopoietic cells among hepatoblasts, and the hepatoblasts made contact with one another by short cytoplasmic projections. The area of the contact surface had a diameter of 4-5 microns, where E-cadherin-mediated adherens junctions were found. At 12-13 days of gestation, hepatoblasts surrounded large ellipsoidal hematopoietic foci, with long cytoplasmic projections. In addition to the adherens junctions, small desmosomes appeared to bind hepatoblasts together, and biliary canaliculi could be recognized between hepatoblasts. At peak stage of liver hematopoiesis at 14 days of gestation, both tight junctions and gap junctions appeared around the biliary canaliculi, and four types of specialized junctions, i.e., adherens junctions, desmosomes, tight junctions and gap junctions, appeared to be fully developed. After 15 days of gestation, hepatocyte volume showed rapid increase, and the surface areas between adjacent hepatocytes were markedly enlarged. As a result, the involuted hematopoietic foci were forced to move from interhepatocytic spaces to perisinusoidal space at the end of the intrauterine life.

Developmental outcomes of children with hearing loss born in Colorado hospitals with and without universal newborn hearing screening programs.

The developmental outcomes of children born in hospitals with universal newborn hearing screening programs were compared with children born in hospitals without universal newborn hearing screening programs. Eight-four percent of children born in screening hospitals were early-identified with hearing loss prior to 6 months of age as compared to 8% of the children in the non-screen group. The participants in the screen group had an average language quotient of 82 while the participants in the non-screen group had an average language quotient of 62. Children in the screen group had better receptive and expressive language quotients, more different consonants in the spontaneous phonetic repertoire, better speech intelligibility, and larger expressive vocabulary inventories. Odds risk ratio estimates indicate that 80% of the children with cognitive quotients 80 or greater or four out of five children had language quotients within the normal range, 80 or greater, when they were in the screen group.

The Colorado Newborn Hearing Screening Project: effects on speech and language development for children with hearing loss.

OBJECTIVE: To compare the speech and language development of children with bilateral hearing loss and normal cognition who were born in hospitals with universal newborn hearing screening to that of their peers who were born in hospitals without this screening program. STUDY DESIGN: Subjects for the major analyses are 50 Colorado children (25 matched pairs) from 9 to 61 months old who are participants in a study of the development of children birth to six with bilateral hearing loss. Analyses included parametric dependent t-tests and analysis of covariance, nonparametric chi-squared and Wilcoxon signed rank tests, descriptive statistics and odds ratios. RESULTS: Newborn screening programs for hearing loss are positively related to scores in expressive and receptive language (p < 0.001) and vocabulary production (p < 0.001) on standardized inventories; speech intelligibility (p = 0.015) from independent ratings; number of different simple consonants (p < 0.01) and consonant blends (p = 0.026) from phonological transcripts; and total number of intelligible words (p < 0.01) and number of different words produced (p < 0.01) from computer analysis of videotaped language samples. CONCLUSION: Hospital-based newborn hearing screening programs are positively related to language and speech performance for children in early intervention programs who are deaf and hard of hearing.

Vimentin intermediate filament protein as differentiation marker of optic vesicle epithelium in the chick embryo.

For the study of the differentiation process of optic vesicle epithelium into neural retina, pigment epithelium and pars caeca retinae, vimentin intermediate filament protein in retinal epithelial cells was detected immunohistochemically in chick embryo at stages 11-21. In the late stage of optic vesicle development (stage 14), optic vesicle epithelium was classified into the following 3 different portions on the basis of vimentin staining intensity: latero-central epithelium under the lens placode, medio-central epithelium facing the latero-central epithelium, and peripheral epithelium connecting the latero-central and medio-central epithelia. Latero-central epithelium, the future neural retina, exhibited strongest staining of vimentin of the 3 portions. In contrast, medio-central epithelium, the future pigment epithelium, showed weakest staining. Moderate staining was observed in peripheral epithelium, the future pars caeca retinae. These differences in levels of vimentin expression were observed during optic cup formation. The present results clearly demonstrate that differentiation of retinal epithelium into neural retina, pigment epithelium and pars caeca retinae occurs in the late stage of the optic vesicle, and that retinal differentiation is reflected by the amount of vimentin in epithelial cells.

Benefits of early intervention for children with hearing loss.

This article discusses the relationship between language development and degree of hearing loss and how these factors relate to the age of identification of hearing loss. An interesting effect caused by the interaction of these variables has been discussed in this article. The relationship of speech production, degree of hearing loss, language development, and age of identification are also presented. Social-emotional development of deaf and hard-of-hearing children and the variables that impact this development, in particular the strong relationship with language development, is included.

The role of educators of the deaf in the early identification of hearing loss.

About 16,000 babies each year will be identified with hearing loss by age 3 months once universal newborn hearing screening becomes a reality. Identification of hearing loss in infancy, followed by appropriate intervention by age 6 months, can result in normal language development, regardless of degree of hearing loss. As the average age of identification of hearing loss moves downward toward 2 months, children with hearing loss will enter the educational system earlier and with language skills commensurate with those of their hearing peers. In order to provide appropriate services to children with hearing loss and their families, early interventionists will need to forge links to health care providers involved in universal newborn hearing screening programs, to have specialized training in deafness and hearing loss, and to have expertise in providing services to very young children and to children with hearing loss in the broad range from mild to profound.

The efficacy of early identification and intervention for children with hearing impairment.

From these findings, the inevitable conclusion is that identification of hearing loss by 6 months of age, followed by appropriate intervention, is the most effective strategy for the normal development of language in infants and toddlers with hearing loss. Identification of hearing loss by 6 months can only be accomplished through universal newborn hearing screening. Some questions that arise as a result of these studies include: What can one conclude from the finding that the language skills of children with mild hearing losses are no better than those with greater losses? If the finding holds up, it indicates a great need for investigations into biobehavior theories of language acquisition and into the part played by the prenatal 4 months of hearing. And it also shows a need for answering the question, When does a hearing loss begin?, because it certainly seems that all hearing losses are similar in their outcomes. Can the findings from these studies be used to benefit normally hearing children who are at risk for language delays as a result of limited language environments? Such children suffer from auditory deprivation just as surely as those with hearing losses. If the language skills of the latter children can be brought to normal range by early intervention, the same strategy may help high-risk populations. The efficacy of early intervention is just as valid for these children as it is for the children with hearing impairment. Now that the benefits of early identification of children with congenital hearing loss have been demonstrated, these benefits should be extended to all children who are at risk for language delays, with appropriate interventions applied immediately.

Maternal sensitivity predicts language gain in preschool children who are deaf and hard of hearing.

The link between maternal sensitivity and child language gain was assessed in a prospective study of hearing mothers and their deaf and hard-of-hearing (D/HH) children. Maternal sensitivity in dyadic interaction was assessed when children were approximately 2 years old, and expressive language gain was assessed at 2 to 3 years using the Minnesota Child Development Inventory. Sensitivity made significant, positive, and unique predictions of expressive language gain when the effects of maternal education, degree of child hearing loss, dyadic mode of communication, and time between assessments were controlled. Findings indicate the value of affective measures of interaction in predicting language gain.

Language of early- and later-identified children with hearing loss.

OBJECTIVE: To compare the language abilities of earlier- and later-identified deaf and hard-of-hearing children. METHOD: We compared the receptive and expressive language abilities of 72 deaf or hard-of-hearing children whose hearing losses were identified by 6 months of age with 78 children whose hearing losses were identified after the age of 6 months. All of the children received early intervention services within an average of 2 months after identification. The participants' receptive and expressive language abilities were measured using the Minnesota Child Development Inventory. RESULTS: Children whose hearing losses were identified by 6 months of age demonstrated significantly better language scores than children identified after 6 months of age. For children with normal cognitive abilities, this language advantage was found across all test ages, communication modes, degrees of hearing loss, and socioeconomic strata. It also was independent of gender, minority status, and the presence or absence of additional disabilities. CONCLUSIONS: Significantly better language development was associated with early identification of hearing loss and early intervention. There was no significant difference between the earlier- and later-identified groups on several variables frequently associated with language ability in deaf and hard-of-hearing children. Thus, the variable on which the two groups differed (age of identification and intervention) must be considered a potential explanation for the language advantage documented in the earlier-identified group.

Effects of colchicine on the enucleation of erythroid cells and macrophages in the liver of mouse embryos: ultrastructural and three-dimensional studies.

Enucleation is the last event in the development of a definitive erythroid line, and extruded nuclei are phagocytosed by macrophages. Both colchicine and cytochalasin have been known to exert a great influence on the enucleation process, but the relationship between enucleation and these agents has not yet been clearly revealed in vivo. Our aim was to clarify the significance of the enucleation in liver erythropoiesis and macrophage phagocytosis by colchicine and cytochalasin administration to embryonic mice. Pregnant mice were intraperitoneally injected with colchicine or cytochalasin at 13 days of gestation. Embryonic livers were removed at intervals of 3, 6 and 12 h after injection for processing for light and electron microscopy, and, to obtain three-dimensional morphology of erythroids at enucleation, computer-aided reconstructions were performed by light microscopy. Colchicine injections had cytolytic effects on hepatocytes and macrophages, and numerous erythroblasts were observed in the process of enucleation after colchicine injection. However, the extruding nuclei were irregularly shaped, and some erythroblasts at mitosis showed extreme peripheralization of their chromosomal masses and cell membrane constriction. Enucleation behavior could also be observed in immature erythroblasts. Liver macrophages engulfed extruded nuclei and erythroblasts in mitosis. Cytochalasin injections, on the other hand, had no significant effect on embryonic livers. The progress of erythroblast mitosis was clearly stopped by colchicine injection, and numerous erythroblasts at mitosis were extruding their nuclear compartment. Following colchicine injection, erythroid enucleation also took place in immature erythroblasts, and mitotic erythroids were phagocytosed. In enucleation, more attention should be paid to hematopoietic environmental factors than to hemopoietic cell factors.

Identification of hearing loss after age 18 months is not early enough.

The general development of 40 deaf and hard of hearing infants was analyzed. The infants were placed into one of two groups according to age at which hearing loss was identified: (a) before age 6 months and (b) after age 18 months. The mean age at testing was 40 months. Developmental quotients (DQs) were used to compare results regardless of the infants' age at time of testing. Infants were evaluated on the basis of their DQ scores on the Minnesota Child Development Inventory (MCDI; Ireton & Thwing, 1972). MCDI subtests include general development, gross motor, fine-motor, expressive language, comprehension-conceptual, situation-comprehension, self-help, and personal-social. Infants whose hearing loss was identified before age 6 months scored significantly higher than those whose hearing loss was identified after age 18 months in the expressive language and comprehension-conceptual subtests. The performance of the earlier-identified group supports the earliest identification of hearing loss and encourages implementation of universal hearing screening.

The development of deaf and hard of hearing children identified early through the high-risk registry.

The high-risk registry was used as a screening device for identifying hearing loss for many decades in Colorado. It reportedly missed approximately 50% of all infants with congenital sensorineural hearing loss (Mehl & Thomson, 1998; Parving, 1993; Watkins, Baldwin, & McEnery, 1991). Little is known about the developmental characteristics of this population. This article describes children identified through the high-risk registry. These children have been divided into two groups according to their age of identification: (a) deaf and hard of hearing children identified before age 6 months, and (b) deaf and hard of hearing children identified between ages 7 and 18 months. The children identified before age 6 months and receiving intervention at an average of 2 to 3 months after identification of hearing loss had significantly higher levels of receptive and expressive language, personal-social development, expressive and receptive vocabulary, general development, situation comprehension, and vowel production. The high-risk registry used for newborn hearing screening has been replaced by universal newborn physiological hearing screening in the state of Colorado.

Origin of the central cells of erythroblastic islands in fetal mouse liver: ultrahistochemical studies of membrane-bound glycoconjugates.

To clarify the origin of the central cells in hepatic erythroblastic islands, glycoconjugates on the surface of cellular constituents in fetal mice liver were ultrahistochemically examined using lectin staining. At 11 days of gestation, the cells derived from mesenchyme in fetal liver, including sinusoidal macrophages, endothelial cells, and erythropoietic cells, bound Griffonia simplicifolia isoagglutinin I-B4 (GS-I-B4), but hepatocytes lacked binding sites for the isolectin. Scavenger macrophages in the hepatic cords at 13 days of gestation and the central cells in the erythroblastic islands at 15 days of gestation also bound GS-I-B4. Hepatocytes, however, exhibited no GS-I-B4 binding site at any gestational day. At 11 days of gestation, none of the cells in fetal liver had binding sites for soybean agglutinin (SBA), but cells derived from mesenchyme acquired these binding sites at 13 days of gestation. The central cells in the erythroblastic islands also bound SBA, but hepatocytes did not bind the lectin at all. The central cells in the erythroblastic islands can be considered to belong to a mesenchymal cell lineage, and primitive sinusoidal macrophages at 11 days of gestation are possible precursors of these central cells.

Death process of primitive erythrocytes and phagocytosis by liver macrophages of the mouse embryo.

Primitive erythrocytes from the yolk sac are nucleated cells which have a short life span of several days in the embryonic circulation of the mouse. The death process of primitive erythrocytes was ultrastructurally investigated in embryonic mice. At 12 days of gestation, primitive erythrocytes accounted for 96.1% of the circulating erythrocytes. The percentages in 13-, 14-, 15- and 18-day-embryos were 43.8%, 15.4%, 7.7% and 0.0%, respectively. Between 13 and 15 days, anucleate primitive erythrocytes made up from 1.5% to 5.9% of circulating erythrocytes. During the gestational period, the nuclei of primitive erythrocytes markedly decreased in volume, and the chromatin underwent condensation and marginated to crescents along the nuclear envelope. Following nuclear fragmentation and dissolution of condensed chromatin, nuclear residues were finally formed in aging primitive erythrocytes. Formation of annulate lamellae was also observed, associated with the nuclear shrinkage. The nuclei had an apoptotic-like appearance, but TUNEL-positive reactions could not be identified in any nuclei of the primitive erythrocytes. Dying erythrocytes with nuclear residues were phagocytosed in toto by hepatic macrophages and disposed of from embryonic circulation without enucleation.

[Scanning and transmission microscopic observations on circulating primitive erythroblasts of yolk sac origin in the mouse embryo].

The earliest hemopoietic tissues which appear during the ontogeny of mammals are the blood islands of the yolk sac, and the blood cells produced therein begin to circulate between the embryo and visceral yolk sac at the establishment of the circulatory system. Primitive erythroblasts derived from the yolk sac have a short life span of only several days, and they form a majority of the embryonic blood cells prior to the start of liver hemopoiesis. To clarify cell fragmentation of primitive erythroblasts at the ultrastructural level, using 18 embryos of ICR-mice at 10 and 11 days of gestation, we observed circulating erythroblasts by scanning and transmission electron microscopy. The circulating erythroblasts generally had an irregularly ovoid contour, and they showed a great deal of micropinocytosis on their cell surface. The nuclei of the erythroblasts were round and possessed one or two nucleoli which were in contact with the nuclear membrane. Their nuclear chromatin was dispersed, and the cytoplasm was rich in polyribosomes and mitochondria. The majority of circulating erythroblasts were at the stage of either basophilic or polychromatophilic erythroblasts. Cytoplasmic projections could occasionally be seen extending from the erythroblast surface, and some of the projections appeared to be liberated into the vascular lumen as cell fragments. On the basis of their size and shape, the cytoplasmic projections could be classified into three types; finger-like projections, vesicular projections and microvesicular projections. The finger-like projections were approximately 1 micron in diameter and 3 microns in length. The vesicular projections, connected with the cell by a narrow stalk, were teardrop in shape, and approximately 0.8 microns in diameter and 1.5 microns in length. The microvesicular projections were approximately 0.2 microns in diameter and 0.2-0.5 microns in length. The finger-like projections had micropinocytotic invaginations on their surface, but no invaginations could be seen on the vesicular and microvesicular projections. Not only the finger-like but also the vesicular projections contained cytoplasmic matrix with a few polyribosomes. The microvesicular projections, on the other hand, occasionally contained myelinic-like figures. These projections were seen on the surface of erythroblasts at various maturation stages. The cytoplasmic fragments released from the erythroblasts were engulfed and eliminated from the embryonic peripheral blood by intravascular macrophages. The fragmentation of cytoplasmic projections was considered to be related to the development of microfilaments involved in the cytoskeleton of the erythroid elements.

The ultrastructure of intravascular and coelomic scavenger macrophages of the mouse embryo.

The aim of this study was to establish whether or not mononuclear cells which appear in both the vitelline vessels and embryonic coelom in mice prior to liver hemopoiesis are specialized scavengers. Before the initiation of liver hemopoiesis, the majority of the embryonic blood cells were primitive erythroblasts derived from yolk sac hemopoietic foci. In addition, the peripheral blood contained a few free phagocytes as early as 10 days of gestation. The phagocytes devoured various cell elements such as degenerating erythroblasts and cell fragments. Ultrastructurally, they had long filamentous cytoplasmic projections on their cell surface, clear subsurface vacuoles or vesicles, lipid droplets, a few lysosomal granules, large heterogeneous phagolysosomes and residual bodies. Mononuclear phagocytes with ultrastructural features similar to those of the intravascular phagocytes also could be observed in the intraembryonic peritoneal cavities at 10 days of gestation; they sometimes engulfed possible mesothelial cells undergoing degeneration. Based on fine structural criteria, these intravascular and coelomic phagocytes were considered to be specialized scavenger macrophages with the function of clearing the blood and tissue fluid of whatever has been ingested. In so doing, they serve as the most primitive discriminating filter set in embryonic circulation.

Membrane-bound glycoconjugates of fetal mouse erythropoietic cells with special reference to phagocytosis by hepatic macrophages.

Using lectin and colloidal iron (CI) stainings in combination with neuraminidase digestion, glycoconjugates on the surface of erythropoietic cells of the yolk sac and liver in fetal mice were examined. Fetal hepatic macrophages were capable of distinguishing between phagocytozed and non-phagocytozed erythroid elements as described in our previous study. Marked differences between these two elements could be ultrahistochemically detected on their cell surface. The phagocytozed elements, such as nuclei expelled from erythroblasts and degenerating primitive erythroblasts, faintly bound neuraminidase-sensitive CI, and neuraminidase digestion imparted a weak peanut agglutinin (PNA) binding. In contrast, erythroblasts at various maturation stages, erythrocytes and normal primitive erythroblasts heavily bound neuraminidase-sensitive CI, and neuraminidase digestion imparted a moderate PNA binding. No differences in binding of either concanavalin agglutinin, Ricinus communis agglutinin-I or PNA were noted between phagocytozed and non-phagocytozed erythroid elements. Desialylation appears to be one of the most important signs for the recognition mechanism of fetal macrophage phagocytosis. During maturation of hepatic erythroblasts, sialic acid changes its affinity for Limax flavus agglutinin from strong to weak, and soybean agglutinin binding sites disappear at the basophilic erythroblast stage. Glycoconjugates on polychromatophilic erythroblasts acquire similar compositions to those of erythrocytes.

Accumulation and massive cell death of polymorphonuclear neutrophils in the developing bone marrow of the mouse: a histological study.

Accumulation and cell death of neutrophils were studied by light and electron microscopy in neonatal mouse bone marrow. At the beginning of bone marrow hematopoiesis, the marrow cavity contained a large number of polymorphonuclear leukocytes. Polymorphs comprised approximately 75% of the total nucleated cells in the hematopoietic compartment of the newborn marrow, the majority being neutrophils. Mature neutrophils were sometimes crossing the endothelium of the marrow blood sinus. Neutrophils in neonatal marrow show features typical of apoptosis, e.g. formation of nuclear pockets and blebs, margination of compact nuclear chromatin to form sharply circumscribed masses, condensation of cytoplasm, and convolution of cell outlines. Dying neutrophils were devoured and digested by phagocytes. The occurrence of large-scale neutrophil death and removal of neutrophils by phagocytes in neonatal bone marrow are discussed in relation to programmed cell death in development of the fetal hematopoietic system.

Cell differentiation of alveolar epithelium in the developing rat lung: ultrahistochemical studies of glycoconjugates on the epithelial cell surface.

Glycoconjugates on the surface of pulmonary epithelial cells were ultrahistochemically examined in the fetal, neonatal and adult rat lung. Lectin and colloidal iron staining procedures were performed in combination with digestion using carbohydrate-degrading enzymes or methylation. The glycoconjugate composition of columnar cells at 16 days gestation was similar to that of cuboidal cells at 19 days gestation. Glycoconjugate differentiation on the cell surface occurred at 20 days gestation, and especially the loss of soybean agglutinin (SBA) binding sites could be detected on type II cells. The contents of Ricinus communis agglutinin-I (RCA-I) and Concanavalin A (Con A) binding sites on type II cells also began to decrease. On the contrary, the content of sulfated saccharides decreased on the surface of type I cells during development. Glycoconjugate differentiation on both type I and II cells was completed with the disappearance of hyaluronic acid and peanut agglutinin (PNA) binding sites; type I and II cells acquired a similar histochemical composition to that on adult type I and II cells at 5 days after birth. Both type I and II cells share a common early precursor cell, that is, the cuboidal epithelial cell at the canalicular stage.