The vit gene maps to the mi (microphthalmia) locus of the laboratory mouse.

The murine model for human vitiligo (the vit/vit mouse) develops progressive depigmentation of the pelage, skin, and eyes. The vit gene is inherited as an autosomal recessive. We have used classical breeding and isozyme marker analysis to map this vit gene that produces a vitiligo-like condition in the mouse. Crossbreeding the C57BL/6J-vit/vit mice with C57BL/6J mice carrying the Miwh and/or miws alleles at the microphthalmia locus resulted in mutant phenotypes, demonstrating absence of complementation. When vit is heterozygous with the Miwh allele, a "blotched" pigment pattern results. When it is heterozygous with the miws allele, a novel expression of the vitiliginous phenotype results. Further mating analysis of these crossbred populations demonstrates allelic inheritance between vit and the alleles at the microphthalmia locus. Other breeding studies using alleles at the agouti, belted, brown, dominant spotting, extension, mahogany, patch, and piebald loci did not demonstrate pigmentation explainable by allelic inheritance with the vit gene. Also, vit was tested for linkage with isozyme markers located on chromosomes 1, 4, 5, 7, 9, and 11, and results were negative. Therefore, the vit (vitiligo) gene of the laboratory mouse has been mapped to the mi (microphthalmia) locus on chromosome 6. The gene properly should be designated as mivit.     

Bone surface structure in osteopetrotic grey lethal (gl/gl) and microphthalmic (mi/mi) mutant mice as revealed by scanning electron microscopy

The surface structure of bone from two genetically distinct osteopetrotic strains of mice, grey lethal (gl) and microphthalmic (mi) has been examined by scanning electron microscopy. Although both conditions produce a classical osteopetrolic phenotype the means by which this is achieved is quite different. gl mice appear to retain woven bone and show no evidence of resorption: mi mice show evidence of imperfect remodelling. These results are in accordance with what is already known of osteoclast structure and function in these mutations.     

Natural killer cells, bone, and the bone marrow: studies in estrogen-treated mice and in congenitally osteopetrotic (mi/mi) mice.

Mice lose natural killer cells after 6 weeks of treatment with 17 beta-estradiol. We here demonstrate that the same protocol leads to loss of genetic resistance to bone marrow transplantation and to significant osteoproliferation with loss of bone marrow. We also show that mice with reduced marrow because of congenital osteopetrosis are deficient in natural killing. These findings are consistent with previous evidence that natural killing and genetic resistance to bone marrow transplantation are dependent upon the marrow. Temporal studies of bone histology and radiology during and after treatment with estrogen reveal that alterations in natural killing proceed more rapidly than changes in bone marrow volume. These studies also demonstrate that estrogens induce osteoproliferation only at endosteal surfaces that are adjacent to hematopoietic marrow. From these observations, we conclude that estrogens do not reduce natural killer cells simply by reducing the volume of bone marrow. Estrogens may instead have an effect on bone marrow. Estrogens may instead have an effect on bone marrow cells that leads both to osteoproliferation and to a deficiency of marrow-dependent cells.     

Morphology of congenital microphthalmia in chicks (Gallus gallus)

This study examines the morphology of sporadic congenital microphthalmia in 1-day-old chicks, with particular emphasis on the neural retina. On the basis of the size of the eyeball it is possible to classify microphthalmia into two groups, severe and mild. In severe microphthalmia (less than 5 mm in equatorial diameter), the eyeball is severely malformed, but in most cases it shows evidence of an organized neural retina. Although ganglion cells and an optic nerve head are present in a small proportion of these retinae, we could not trace an optic nerve projection to the brain. These results indicate that some ganglion cells are able to be sustained after the period of naturally occurring cell death, suggesting either that those ganglion cells have established some contact with the central nervous system or that the presence of their axons in a rudimentary optic nerve is adequate for survival. In mild microphthalmia (greater than 5 mm in equatorial diameter), the most consistent abnormality is a defect in the pecten, which together with other abnormalities such as orbital cysts and colobomas indicates that the major abnormality occurs in the region of the choroid fissure. Associated with these defects are abnormalities within the ganglion cell layer. In some cases the number of ganglion cells was reduced, and in others the numbers of both ganglion and displaced amacrine cells were reduced. Unexpectedly, there were localized regions completely devoid of cells in the ganglion cell layer. The timing of the congenital defect may provide some clue as to the presence of a critical period in which displaced amacrine cells are formed or are sensitive to events related to ganglion cell loss.     

Role of microphthalmia transcription factor (Mitf) in melanoma differentiation

We transfected the melanocyte-specific Mitf-M isoform into the aggressive melanoma UISO-Mel-6 cell lines. Our data show that Mitf decreases cell proliferation and results in cells which grow in clusters. By analyzing the expression of the markers of differentiation, we demonstrate that Mitf favored increased expression of tyrosinase and tyrosinase-related protein-1. In addition, Mitf induces Bcl-2 expression following transfection of UISO-Mel-6 cells. We also showed that Mitf gene affects cell-cycle distribution by resting cells preferentially in G2/G1 phase, and inducing the expression of p21 and p27. Moreover, we performed in vivo studies using subcutaneous injection of UISO-Mel-6 and UISO-Mel-6-Mitf in Balb/c nude mice. Our data show that Mitf inhibits tumor growth and decreases Ki67 expression. Tumors induced by UISO-Mel-6 cells were ulcerated and resulted in metastases to liver. None of the mice injected with UISO-Mel-6(Mitf+) cells harbored liver metastases. Our results suggest that Mitf is involved in melanoma differentiation and leads to a less aggressive phenotype.     

Mast cells in spotted mutant mice (W Ph mi)

Mast cells derived from haematopoietic tissue are deficient in numbers in spleen, stomach and skin of Harwell mice doubly mutant at the spotting W locus: seven viable combinations of four mutants. Most combinations have variably impaired viability, anaemia and infertility; but homozygous WshWsh are normal in these respects yet still lack mast cells. The effect of the W gene on mast cells acts in recessive fashion. Effects of doubly mutant W genes on mast cells and coat colour, the latter usually regarded as dominant, appear more closely related than other pleiotropic effects. The spotting gene Ph, closely linked to W, has but marginal effects on mast cells, whereas mi, another spotting gene, quite unrelated to W affects mast cells in the spleen in a dominant way. Thus, splenic mast cells may be a special category of a heterogeneous population. Peptic ulceration, recorded in W/Wv mice of Jackson stock, was not seen in Harwell mice. We suggest that this lesion is due to genetic complementation or environmental causes.     

Comparison of bone and parathyroid hormone as stimulators of osteoclast development and activity in calvarial cell cultures from normal and osteopetrotic (mi/mi) mice

Osteoclast development was studied in cell cultures prepared from calvaria of neonatal osteopetrotic (mi/mi) mice or their normal littermates, using tartrate-resistant acid phosphatase (TRAPase), as an osteoclast marker. In cultures from normal mice, treatment with 10 nM PTH for 4-5 days stimulated the formation of osteoclasts. However in cultures from mi/mi mice, this response was only 7% +/- 5% that of normal mice and they were significantly smaller than osteoclasts of normal mice. Mineralized bone particles elicited osteoclast development in cultures from both normal and mi/mi mice, and osteoclast size was identical for both genotypes. Seventy-eight to 96% of the TRAPase-positive cells bound 125I-CT, as demonstrated by autoradiography. 125I-CT binding characteristics were identical in cultures from both genotypes treated with bone particles, exhibiting a Kd of 3.3-3.6 x 10(-10) M. Addition of PTH stimulated 45Ca release from the added bone particles only in the case of cultures prepared from normal mice, and CT inhibited this response. Cells from normal mice were capable of excavating bone from the surface of smooth cortical bone wafers, but such excavations were rarely seen in the case of calvarial cells from mi/mi mice. Thus, PTH-driven differentiation of osteoclasts is arrested in calvarial cell cultures from mi/mi mice, but mi/mi preosteoclasts retain the ability to express certain osteoclast markers in response to bone derived signals. We hypothesize that the lack of activity of mi/mi osteoclasts is due to the failure of mi/mi preosteoclasts to respond appropriately to resorptive agents, or to cytokines elicited by these agents.     

Morphometric analysis of the dorsal lateral geniculate body of the laboratory mouse and microphthalmic variants

The dorsal lateral geniculate nucleus (dLGN) of the mouse (strain 944 with microphthalmus syndrome, P. Hertwig 1942) has been morphometrically investigated. The eyes of the gray littermates develop normally (control animals) while the white littermates show the microphthalmus syndrome (microphthalmic animals). Four male twentyone day old animals were examined. Following perfusion with formaldehyde (diluted 1:4 with H2O) Nissl stained frontal sections were used for classification of neurons. we differentiate between projection neurons (PN) (geniculo-cortical relay neurons) and interneurons (IN). We succeeded in identifying these two cell types using deimpregnated Golgi preparations (Fairén et al. 1977), which were counterstained with methylene blue (0.1%). The neuronal packing density and the ratio PN:IN were conventionally determined using 20 micron thick sections from both control and microphthalmic animals. In both cases this ratio was 12:1 (= 8% IN). Measuring fields distributed in a meander-like manner were selected for estimating the neuronal density. Measuring fields giving equal values were connected. The results show that we can distinguish between 3 regions of dLGN: lateral, medial and caudal. They differ with regard to neuronal packing density, size and structure of the PN. Using 3 micron thick Nissl stained sections from both control and microphthalmic animals the size and structure of PN were automatically determined using the picture processing device "MORPHOQUANT" VEB Carl Zeiss JENA). The distribution pattern of IN is apparently uniform. The microphthalmus syndrome produces a diminished number of pathologically changed retinal fibers. The morphometrically registered findings show that within the dLGN, of the microphthalmus mouse the neuronal packing density increased (diminution of neuropil), the size of PN decreased and their structural density increased (transneuronal dystrophy). Compared with the other parts, the medial part of the dLGN is minimally altered.     

The hematogenous origin of osteoclasts: experimental evidence from osteopetrotic (microphthalmic) mice treated with spleen cells from beige mouse donors

The excessive skeletal mass and reduced bone resorption characteristic of osteopetrosis in microphthalmic (mi) mice can be corrected by irradiation and transfer of spleen cells from a normal littermate. Osteoclasts in beige (bg) mice, a mutation without osteopetrosis, have giant lysosomal granules. These two facts were exploited to trace osteoclast lineage. Microphthalmic mice treated with whole-body irradiation and spleen cells from a beige donor resorbed the excessive skeletal mass and recovered from osteopetrosis. Furthermore, osteoclasts in treated mi mice had giant lysosomal granules and resembled those found in bg donors when examined by light and transmission electron microscopy. These data provide direct evidence for a hematogenous origin of osteoclasts in mammals.     

Association between the lack of teeth and the expression of myosins in masticatory muscles of microphthalmic mouse

The purposes of the present study were to elucidate the influences of the deficiency of teeth on masticatory muscles, such as the masseter, temporalis and digastric muscles and compare the influence among masticatory muscles. We analysed the expressions of myosin heavy chain (MyHC) isoform messenger RNA (mRNA) and protein in these muscles in the microphthalmic (mi/mi) mouse, whose teeth cannot erupt because of a mutation in the mitf gene locus. The expression levels of MyHC mRNA and protein in the masseter, temporalis, digastric, tibialis anterior and gastrocnemius muscles of +/+ and mi/mi mice were analysed with real‐time polymerase chain reaction and sodium dodecyl sulfate‐polyacrylamide gel electrophoresis, respectively. The mi/mi masseter muscle at 8 weeks of age expressed 4·1‐fold (p < 0·05) and 3.3‐fold (p < 0·01) more MyHC neonatal mRNA and protein than that in the +/+, respectively; the expression level of MyHC neonatal protein was 19% of the total MyHC protein in the masseter muscle of mi/mi mice. In the digastric muscle, the expression levels of MyHC I mRNA and protein in the mi/mi mice were 4·7‐fold (p < 0·05) and 5‐fold (p < 0·01) higher than those in the +/+ mice. In the temporalis, tibialis anterior and gastrocnemius muscles, there was no significant difference in the expression levels of any MyHC isoform mRNA and protein between +/+ and mi/mi mice. These results indicate associations between the lack of teeth and the expression of MyHC in the masseter and digastric muscles but not such associations in the temporalis muscle, suggesting that the influence of tooth deficiency varies among the masticatory muscles. Copyright © 2011 John Wiley & Sons, Ltd.     

Defect of a fiber cell-specific 94-kDa protein in the lens of inherited microphthalmic mutant mouse Elo

Deficiency in a 94,000-dalton protein in the non-crystallin fraction from the Elo mouse lens was shown. To perform further investigations, we raised an antibody against the 94,000-dalton protein isolated from normal mouse lens. Western blot analysis with the antibody indicated that the protein was only present in the lens and not in the brain, lung, heart, liver, and kidney. In the lens, it was unique to the cortex and nucleus fractions, not being present in the epithelial cells. Furthermore, it was observed in the water-soluble fraction as well as in the urea-soluble fraction. The antibody weakly but clearly reacted with the chick CP97 lens peptide, a fiber cell-specific protein, and anti-CP97 antibody also reacted with the 94,000-dalton protein. From these results, we concluded that the protein corresponds to CP97 cytoskeletal protein in the mouse lens. The protein was deficient in the lenses from Elo mice, but microphthalmic lenses from CTA mice contained a normal level.     

Microphthalmia (mi) mice display an aberrant bone trace element composition

It has been recognized that bone trace element composition analysis provides clues when analyzing bone-related physiological conditions. Increasing numbers of bone-related genetic diseases have been identified recently. In this study, we have analyzed bone trace element composition in a genetic mutant animal model. Mutations in the mouse microphthalmia (mi) gene affect the development of a number of cell types, including melanocytes, mast cells, and osteoclasts. Previous studies have shown that different alleles of the mi locus show osteopetrosis. In order to gain insights into the effects of a particular genetic defect on bone trace element composition and bone structure, we performed bone trace element composition analysis using inductively coupled plasma atomic emissions spectrometry (ICP-AES). Marked changes in bone trace element levels were found in vertebrate bones of mi mutant mice. The implications and possible applications of bone trace element analysis will be discussed in this article.