Biomedical subjects

L Mirels

Publications and source records attributed to L Mirels.

At least 19 recordsLinked to original sources

PubMed2003

Psp and Smgb: a model for developmental and functional regulation in the rat major salivary glands.

This paper summarizes past work detailing the developmental expression, cell and organ localization and biochemical features of the proteins parotid secretory protein (PSP) and isoforms of submandibular gland protein B (SMGB), and describes the molecular characterization of the genes that encode them, Psp and Smgb. These genes appear to be related to the BPI (bactericidal/permeability-increasing protein)/LBP (lipopolysaccharide-binding protein)/PLUNC (palate, lung and nasal epithelial clone) gene family found in the oral and respiratory organs of humans, rodents and cattle. We have emphasized the diverse patterns of expression of these genes among the submandibular, sublingual and parotid salivary glands of the rat, and their potential usefulness in defining and identifying genomic regulatory mechanisms of salivary development. While Psp is expressed similarly in the mouse, the putative Smgb gene of the mouse seems not to be expressed, apparently due to the insertion, between exons 1 and 2, of a gene for a retroviral protein.

Animals↗

PubMed2002

Development of the rat sublingual gland: a light and electron microscopic immunocytochemical study.

Cell differentiation in the rat sublingual gland occurs rapidly and is largely complete by birth. To study differentiation of the serous and mucous cells of the sublingual gland, we used antibodies to the secretory proteins CSP-1, SMGB, PSP, and SMGD, and sublingual mucin as specific cell markers. Glands from rats at ages 18, 19, and 20 days in utero, and postnatal days 0, 1, 5, 9, 14, 18, 25, 40, and 60 were fixed and prepared for morphological analysis and immunocytochemical labeling. At age 18 days in utero, a few cells in the developing terminal bulbs contained mucous-like apical granules that labeled with anti-mucin. Other cells had mixed granules with a peripheral lucent region and a dense core of variable size that occasionally labeled with anti-SMGD. Additionally, presumptive serous cells with small dense granules that contained CSP-1 and SMGB were present. At age 19 days in utero, the dense granules of these cells also labeled with anti-SMGD. By age 20 days in utero, mucous cells were filled with large, pale granules that labeled with anti-mucin, and serous cells had numerous dense granules containing CSP-1, SMGB, PSP, and SMGD. Fewer cells with mixed granules were seen, but dense regions present in some mucous granules (MGs) labeled with anti-SMGD. After birth, fewer MGs had dense regions, and serous cells were organized into well-formed demilunes. Except for PSP, which was undetectable after the fifth postnatal day, the pattern of immunoreactivity observed in glands of neonatal and adult animals was similar to that seen by age 20 days in utero. These results suggest that mucous and serous cells have separate developmental origins, mucous cells differentiate earlier than serous cells, and cells with mixed granules may become mucous cells.

Animals↗

PubMed2000

Structure and chromosomal localization of the rat salivary Psp and Smgb genes.

SMGB and PSP are among the most abundant products of the immature acinar cells in developing rat parotid and submandibular glands and are also products of the sublingual gland serous demilunes. Previous analysis of Smgb and Psp cDNA clones demonstrated a high degree of sequence similarity between the signal peptide-encoding and 3' untranslated regions of these transcripts, although the secreted proteins themselves are more divergent. The current study reports the upstream sequences, genomic organization and localization of the Psp and Smgb genes. Both structural genes contain nine exons and are present at 3q41-3q42, where they are arranged in tandem and separated by 21kb. In addition to the previously observed sequence similarity, Psp and Smgb are highly homologous throughout exon 1 and at 365 of 600bp immediately upstream of the transcription start site. These findings indicate that the Psp and Smgb genes arose by tandem duplication and divergence. The similar neonatal submandibular and parotid gland expression patterns observed for these genes are likely to be due to closely conserved or shared enhancer(s).

Animals↗

PubMed2000

Submandibular gland adenocarcinoma of intercalated duct origin in Smgb-Tag mice.

A line of transgenic mice that develops submandibular gland adenocarcinoma of intercalated duct origin was established. In these mice, the oncogene SV40 T antigen (Tag) is expressed from the neonatal submandibular gland secretory protein b (Smgb) gene promoter. This hybrid gene directs expression of the oncoprotein to neonatal submandibular gland proacinar and terminal tubule cells and to intercalated ducts of the adult gland. Transgene expression resulted in duct luminal cell hyperplasia as early as 20 to 30 days postnatally, which progressed to dysplasia by 3 to 4 months of age. Marked dysplasia and in situ carcinoma were evident at 4 to 6 months of age. All histologic changes were more pronounced in males. Submandibular gland adenocarcinoma developed stochastically in more than half of the adult male mice by 12 months of age (average age: 10.8 months, range: 6 to 13.5 months). Tag expression persisted in in situ carcinoma and all tumors. Using a combination of immunocytochemical and ultrastructural criteria, submandibular gland dysplasia and tumors were found to originate from intercalated ducts. The dysplastic ducts and adenocarcinoma in Smgb-Tag mice were morphologically similar to previously reported Tag-induced dysplasias of striated ducts and granular convoluted tubules and a Tag-induced adenocarcinoma of striated duct origin. These findings demonstrate that salivary gland dysplasias and tumors of similar histologic appearance can arise from distinct differentiated cell types. Analysis of the molecular changes accompanying tumor formation in Smgb-Tag mice could increase knowledge of human salivary gland tumorigenesis.

Adenocarcinoma↗

PubMed1998

Characterization of the rat salivary-gland B1-immunoreactive proteins.

The B1-immunoreactive proteins (B1-IPs) are major secretory products of rat submandibular gland acinar-cell progenitors, and are also produced by neonatal and adult rat sublingual and parotid glands. In order to characterize the B1-IPs, we have previously isolated cDNA clones encoding rat parotid secretory protein (PSP; the predominant parotid B1-IP) and the related clone ZZ3, which is developmentally regulated in the neonatal submandibular gland. The remainder of the B1-IPs were uncharacterized. This report demonstrates that all of the B1-IPs are derived from the PSP and ZZ3 transcripts. Molecular cloning and Western-blot analyses using PSP- and ZZ3-specific antisera show that, of the B1-IPs, only PSP and neonatal submandibular gland protein A (SMGA) are products of the Psp gene. This finding corrects our previous assertion that SMGA is derived from ZZ3. Neonatal submandibular gland proteins B1 and B2, as well as apparent Mr 26000-28000 and Mr 18000-20000 forms in submandibular, sublingual and parotid glands, are derived from the gene encoding ZZ3 by differential N-glycosylation and by proteolytic cleavage. The apparent Mr 18000-20000 proteolytic products are significant in secretion product collected in vitro, but rare in gland homogenate and submandibular/sublingual saliva. The gene encoding ZZ3 has been named Smgb. Psp and Smgb are regulated similarly in the developing submandibular gland, but differently in the sublingual and parotid glands. The expression pattern of Psp is conserved between rat and mouse. However, no evidence for proteins derived from an Smgb-like gene was observed in neonatal mouse submandibular or sublingual glands.

Animals↗

PubMed1998

Secretory protein expression patterns during rat parotid gland development.

The rat parotid gland produces a number of well-characterized secretory proteins. Relatively little is known, however, about the onset of their synthesis and cellular localization during gland development. Secretory protein expression was studied in parotid glands of fetal and postnatal rats using light and electron microscopic immunocytochemistry and Northern blotting. Amylase, parotid secretory protein (PSP), common salivary protein-1 (CSP-1), and SMGB were first detected by immunofluorescence in parotid glands of 18 day fetuses. By 5 days after birth, light and electron microscopic immunolabeling localized all of these proteins to the secretory granules of developing acinar cells. Labeling of acinar cells for DNAse I, however, was not observed until 18 days after birth. Between 9 and 25 days, CSP-1 and SMGB reactivity of acinar cells declined, but increased in intercalated duct cells. After 25 days, CSP-1 and SMGB were found only in intercalated ducts, and amylase, PSP, and DNAse I were restricted to acinar cells. Levels of CSP-1 and SMGB mRNA were relatively constant through 21 postnatal days, but declined significantly after that. Amylase and PSP mRNA increased rapidly and continuously from five days after birth to the adult stage. In contrast, DNAse I mRNA was not detectable until 18 days after birth. The immunocytochemical and molecular analyses define three basic patterns of protein expression in the rat parotid gland: proteins whose synthesis is initiated early in development and is maintained in the acinar cells, such as amylase and PSP; proteins that are initially synthesized by immature acinar cells but are restricted to intercalated ducts in the adult gland, such as CSP-1 and SMGB; and proteins that are synthesized only by mature acinar cells and first appear during the third postnatal week, such as DNAse I. The parotid gland exhibits four distinct developmental stages: prenatal, from initiation of the gland rudiment until birth; neonatal, from 1 day up to about 9 days postnatal; transitional, from 9 days to 25 days of age; and adult, from 25 days on. Although differences exist in timing and in the specific proteins expressed, these developmental stages are similar to those seen in the rat submandibular gland. Additionally, the results support the suggestion that intercalated ducts may differentiate from the neonatal acini.

Age Factors↗

PubMed1998

Expression of gross cystic disease fluid protein-15/Prolactin-inducible protein in rat salivary glands.

Gross cystic disease fluid protein-15 (GCDFP-15)/prolactin-inducible protein (PIP) is present at moderate levels in human submandibular and sublingual glands and is barely detectable in human parotid gland. The rodent homologue, PIP, has previously been identified in adult submandibular and lacrimal glands. Here we present the molecular characterization of rat PIP and show that this protein is a product of neonatal and adult rat submandibular, sublingual, and parotid glands. cDNA clones encoding rat PIP were isolated and sequenced. The deduced amino acid sequence of rat PIP shows 56% overall identity and 80% similarity with mouse PIP. By SDS-PAGE, secreted rat PIP has an apparent Mr of 17,000, with a minor proportion present as Mr 20-22,000 N-glycosylated forms. PIP was localized in rat salivary glands by immunogold silver staining. PIP was identified in acinar cells of developing and mature submandibular and parotid glands and at very low levels in sublingual gland serous demilunes. Typically, rat submandibular gland secretory proteins are produced by either acinar cell progenitors (Type III cells) or mature acinar cells. The expression pattern observed for PIP is similar to that previously reported for salivary peroxidase, an important component of nonimmune mucosal defense.

Aging↗

PubMed1996

Immunocytochemical studies of cell differentiation during rat salivary gland development.

The development of the rat submandibular and parotid glands has been studied using antibodies to secretory proteins as cell-specific markers. Although the morphology of the glands and the timing of the main steps of cytodifferentiation are substantially different, they have several proteins and developmental features in common. The latter include the initial formation of perinatal acini which undergo a transition to adult acini expressing a different complement of secretory proteins; the development of adult intercalated ducts (ID) from the perinatal acinar cells; and the retention of the perinatal phenotype in some cells of the ID.

Animals↗

PubMed1996

Mouse submandibular gland salivary apomucin contains repeated N-glycosylation sites.

A cDNA clone encoding mouse submandibular gland salivary mucin apoprotein was isolated and characterized. The mucin cDNA encodes a protein of 273 amino acids with a calculated molecular weight of 29,606. This apomucin is approximately 60% Thr, Ser, and Pro, and has a pI of 11.25. In addition to the signal sequence, the apomucin can be divided into four structural domains. The first of these contains over 30% Thr, Ser, and Pro, but only a few probable O-glycosylation sites. The second domain contains 10 repeats, each 9 or 13 amino acids in length, with Thr representing more than 50% of the amino acids, while Ser accounts for only 2%. Each repeat begins with a putative N-glycosylation site; hence this domain likely contains both N- and O-linked oligosaccharides. The third domain lacks a repeat motif, but is rich in both Thr and Ser, and therefore is potentially highly O-glycosylated. The final domain is composed mainly of basic and non-polar amino acids and does not contain Cys. This mucin shows considerable homology with the rat submandibular salivary mucin, but little overall homology with other mucins. In situ hybridization verifies that the mucin transcript is localized primarily in the acinar cells of the submandibular gland.

Amino Acid Sequence↗

PubMed1993

Characterization of common salivary protein 1, a product of rat submandibular, sublingual, and parotid glands.

We have isolated and characterized cDNA clones derived from a developmentally regulated neonatal rat submandibular gland salivary protein gene called "common salivary protein 1" (CSP1). Identical clones were also identified in cDNA libraries from adult male parotid, submandibular, and sublingual glands. CSP1 transcripts are at least 10-fold more abundant in the sublingual gland than in the submandibular or parotid glands. In situ hybridization and immunocytochemical localization demonstrated the presence of CSP1 transcripts and proteins in sublingual gland serous demilune cells, parotid and submandibular gland intercalated duct cells, and in the type III (proacinar) cells of the neonatal submandibular gland. This cell-type distribution is similar to that described by Ball and colleagues (Ball, W. D., Hand, A. R., and Johnson, A. O. (1988) Dev. Biol. 125, 265-279) for the developmentally regulated submandibular gland B1-immunoreactive proteins. Immunoblotting of salivary secretion identified proteins of M(r) 20,000 in sublingual, 16,000 in submandibular and 22,000 and 16,000 in parotid gland. The M(r) 20,000 sublingual and 22,000 parotid proteins represent N-glycosylated forms of a M(r) 16,000 apoprotein, suggesting that these salivary proteins arise by post-translational modification of a common precursor.

Amino Acid Sequence↗

PubMed1993

Molecular cloning of developmentally regulated neonatal rat submandibular gland proteins.

At birth, the rat submandibular gland (SMG) contains two transient secretory cell types that produce several characteristic salivary proteins. Proteins SMG-A, B1, and B2 (23.5, 26 and 27.5 kDa) are products of the neonatal type III cells, but not the adult acinar cells. Protein C (89 kDa), a major product of the neonatal type I cells, is either absent or present at greatly diminished levels in the secretory cells of the adult gland. The decrease in biosynthesis of these neonatal salivary proteins occurs concomitantly with the increase in levels of characteristic adult SMG products. In order to understand these developmentally regulated changes in SMG salivary protein gene expression, we have initiated the molecular cloning and characterization of neonatal submandibular gland proteins from a 5-d-old rat submandibular gland cDNA library. Clones encoding SMG-A were isolated by homology to the mouse parotid secretory protein (PSP). SMG-A was shown to be derived from a salivary protein multigene family that also includes PSP. Cloning and characterization of additional neonatal rat submandibular gland proteins was initiated by screening the 5-d-old rat submandibular gland cDNA library with first strand cDNA prepared from 1-d-old rat submandibular glands. Clones corresponding to a highly abundant 3 kb transcript present in the neonatal rat SMG, but not in adult submandibular, sublingual, or parotid gland have been identified. The size, abundance, and organ specificity of this transcript suggest that it may encode protein C.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1992

Neonatal rat submandibular gland protein SMG-A and parotid secretory protein are alternatively regulated members of a salivary protein multigene family.

Submandibular gland-A (SMG-A) is a major secretory product of the neonatal rat submandibular gland but is not synthesized by the acinar cells of the adult gland. The leucine-rich protein is a predominant product of the adult rat parotid gland. Preliminary data had indicated that the leucine-rich protein and SMG-A might be identical proteins whose expression was differently regulated in the parotid and submandibular salivary glands. cDNA clones encoding SMG-A and the leucine-rich protein were identified by homology to a major mouse parotid gland product, parotid secretory protein (PSP), and characterized. The leucine-rich protein shares extensive sequence homology with mouse PSP throughout its 5'-untranslated, protein coding and 3'-untranslated regions, prompting our suggestion that it should henceforth be referred to as rat PSP. SMG-A is more divergent, having greatest identity with rat and mouse PSP in its signal peptide and 3'-untranslated sequences. Transcripts homologous to SMG-A and rat PSP, but more closely related to SMG-A, were identified in rat sublingual gland by Northern blot analysis. These findings indicate that rat SMG-A and PSP arise from alternatively regulated members of a multigene family.

Amino Acid Sequence↗

PubMed1991

Accumulation and localization of two adult acinar cell secretory proteins during development of the rat submandibular gland.

The seromucous acinar cells of the adult rat submandibular gland secrete a characteristic mucin glycoprotein and a family of unusual glutamine/glutamic acid-rich proteins (GRP). Monoclonal antibodies to the mucin and GRP localized in a very few Type III cells in glands of newborn and 1 day-old rats, using light and electron microscopic immunocytochemistry. Both mucin and GRP reactivities were present in the polymorphic Type IIIP granules during the 1st postnatal week. By 9 days after birth, the granules contained both mucin and GRP and were mucous-like in appearance. At earlier stages, however, cells containing only GRP or mucin could be found, indicating that the initiation of GRP and mucin biosynthesis may not be coordinately regulated. No reactivity was seen in the neonatal Type I cells or in duct cells at any age. Northern and Western blot analysis showed GRP mRNA and protein levels to be barely detectable at birth, with marked increases during the first 2 postnatal weeks. In contrast, Western blots of B1-immunoreactive proteins (B1-IP) showed levels highest in the 1st week and markedly decreased in the adult. Immunocytochemical colocalization, using gold particles of different sizes, showed that the B1-IP, mucin, and GRP colocalized in the granules. These results strengthen the hypothesis that the adult acinar cells develop from the neonatal Type III cells. No evidence was obtained for the involvement of Type I cells in the pathway of acinar cell development.

Aging↗

PubMed1990

Expression of glutamine/glutamic acid-rich proteins in rat submandibular glands.

The expression of GRP transcripts was found to be highly specific to the rat submandibular gland. GRP cross-reactive species were detected in the saliva of both inbred and outbred rat strains. There was no evidence of GRP transcripts in RNA prepared from bovine, ovine, porcine or murine submandibular glands. Thus the GRPs differ from the family of PRPs that are expressed in several species. The restriction of GRP expression to the rat suggests a relatively recent origin for a functional GRP gene, presumably after rat-mouse divergence. The ontogeny of the relative steady-state levels of GRP transcripts was assessed by dot blot analysis. Maximal levels of RNA-encoding GRP were detected at 6 months; there was a significant age-related decline at both 12 and 18 months. There was, however, no significant age-related alteration in the size of transcripts which encode this protein family.

Animals↗

PubMed1989

Dissociation between c-fos gene expression and DNA synthesis in rat parotid glands.

Two experimental approaches were used to examine the relationship between c-fos gene expression and tissue proliferative responses. Beta-Adrenergic and muscarinic receptor stimulation yielded equivalent levels of c-fos expression, although only beta-adrenergic receptor agonists are reported as capable of eliciting DNA synthesis in parotid cells. Similarly, beta-adrenergic stimuli evoked comparable levels of c-fos expression in parotid cells from 2- and 12-month-old rats, whereas DNA synthesis has been shown to be much greater in younger animals. The results indicate that enhanced c-fos expression by itself is incapable of eliciting proliferative responses in rat parotid glands.

Aging↗

PubMed1989

Rapid evolution of variants in a rodent multigene family encoding salivary proteins.

A survey of polypeptides encoded by RNA isolated from the submandibular glands of members of the Muridae (species of Mus and Rattus), in conjunction with cDNA cloning, has identified a class of salivary proteins that we term "spot proteins." Although clearly homologous, these proteins show dramatic differences between species in their polypeptide length. On the basis of the sequence of the corresponding clones, it is inferred that the rat spot 1 protein has a size of 6,370 daltons (Da), whereas that of the inbred mouse spot 1 is 11,603 Da. A second component is expressed in some stocks and strains of Mus, and this spot 2 protein has a size of up to 19,212 Da. The sizes of the corresponding mRNAs show parallel differences, and the variation in the sizes of mRNAs in different species of Mus correlates with the pattern of speciation, the size increasing with increased relatedness to inbred mice. The spot protein sequence comprises three domains: an N-terminal domain rich in hydroxy and acidic amino acids, a central domain consisting of repeats of a 9-amino-acid sequence, and a C-terminal domain that in the mouse is very basic. Variation in the number of repeats largely accounts for the differences in size between the mouse and rat mRNAs and their encoded polypeptides, and the coding sequence appears to have been expanding during speciation in the Muridae. There is extensive divergence in sequence between the mouse and rat mRNAs and their encoded proteins. The pattern of amino acid replacements and nucleotide substitutions is consistent with little, if any, selection constraint on the precise sequence of the spot proteins, suggesting that it is the overall architecture of the molecule, rather than the precise structure, that is important for function. There is strong evidence for a gene conversion event having occurred between the two mouse sequences. Frequent recombination by unequal crossing-over between spot protein coding sequences, if it occurs between active and silent genes, could account not only for the expansion in their size but also for their rapid divergence.

Animals↗

PubMed1987

Molecular characterization of glutamic acid/glutamine-rich secretory proteins from rat submandibular glands.

A family of abundant rat submandibular gland secretory proteins has been identified in glandular extracts and characterized. By amino acid analysis these proteins contain approximately 35% glutamic acid and glutamine plus 14% proline. They have therefore been named "Glx-rich proteins" (GRP). Plasmids containing cDNAs for a GRP have been isolated from a cDNA library prepared from rat submandibular gland poly(A)+RNA. The nucleotide sequence of these cDNAs have been determined. Approximately half of the protein coding sequence is composed of a 23-residue tandem repeat which is repeated five times. The first four repeats are highly conserved at both the nucleotide and amino acid level and consist of the prototype sequence: Asn-Gln-Glu-Pro-Pro-Ala-Thr-Ser-Gly-Ser-Glu-Glu-Glu-Gln-Gln-Gln-Gln-Glu- Pro-Thr-Gln-Ala-Glu. The expression of GRP appears to be specific to the submandibular gland. In vitro assays demonstrate that the GRP have a marked affinity for hydroxyapatite. This suggests that GRP may play a role in the formation of the protective acquired pellicle at the saliva-tooth interface.

Amino Acid Sequence↗

PubMed1985

Isolation and characterization of a mucin-glycoprotein from rat submandibular glands.

A blood group A+ mucin-glycoprotein was purified from aqueous extracts of rat submandibular glands by sequential chromatography on columns of Sepharose CL-6B and Sephacryl S-300 in urea-containing buffers. Final purification was facilitated by reductive methylation which appeared to release contaminating (hydrophobic) peptides. Homogeneity of the purified mucin was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis at varying concentrations of acrylamide, lectin affinity chromatography, and Western blot analysis. In contrast to previously described preparations, the purified mucin contained only trace amounts of N-acetylglucosamine and aromatic amino acids. In addition, only low levels of basic amino acids were present.

ABO Blood-Group System↗