Senescence Marker Protein-30 Knockout Mice Research Articles

Immunohistochemical localization of senescence marker protein-30 (SMP30) in the submandibular gland and ultrastructural changes of the granular duct cells in SMP30 knockout mice.

Senescence Marker Protein-30 (SMP30) is a calcium-regulating protein that decreases in an androgen-independent manner as aging occurs. An enzyme-labeled antibody technique has demonstrated that SMP30 localized to the ducts (granular, intercalated, and striated ducts) of mouse submandibular glands. Immunoelectronmicroscopy demonstrated that the granular duct cells were strongly positive for SMP30, but that pillar cells in the granular duct were negative for the protein. In SMP30-knockout (KO) mice, the granular ducts were smaller in diameter. Swelling of mitochondria in the granular duct cells was observed; however, this phenomenon was not observed in the pillar cells. After administration of alpha-isoproterenol, a beta-adrenergic stimulant, a large numbers of small secretory granules were present in the granular duct cells and an expansion of the rough endoplasmic reticulum in SMP30-wild type (WT) mice; in contrast, little change was observed in SMP30-KO mice. These results suggest that SMP30 may be closely related to a signal transduction pathway in the granular duct cells of submandibular glands.

Senescence marker protein-30 is a unique enzyme that hydrolyzes diisopropyl phosphorofluoridate in the liver

Senescence marker protein-30 (SMP30) was originally identified as a novel protein in the rat liver, the expression of which decreases androgen-independently with aging. We have now characterized a unique property of SMP30, the hydrolysis of diisopropyl phosphorofluoridate (DFP), which is similar to the chemical warfare nerve agents sarine, soman and tabun. Hydrolysis of DFP was stimulated equally well by 1 mM MgCl 2, MnCl 2 or CoCl 2, to a lesser extent by 1 mM CdCl 2 but not at all by 1 mM CaCl 2. No 45Ca 2+-binding activity was detected for purified SMP30, suggesting that SMP30 is not a calcium-binding protein, as others previously stated. Despite the sequence similarity between SMP30 and a serum paraoxonase (PON), the inability of SMP30 to hydrolyze PON-specific substrates such as paraoxon, dihydrocoumarin, γ-nonalactone, and δ-dodecanolactone indicate that SMP30 is distinct from the PON family. We previously established SMP30 knockout mice and have now tested DFPase activity in their livers. The livers from wild-type mice contained readily detectable DFPase activity, whereas no such enzyme activity was found in livers from SMP30 knockout mice. Moreover, the hepatocytes of SMP30 knockout mice were far more susceptible to DFP-induced cytotoxicity than those from the wild-type. These results indicate that SMP30 is a unique DFP hydrolyzing enzyme in the liver and has an important detoxification effect on DFP. Consequently, a reduction of SMP30 expression might account for the age-associated deterioration of cellular functions and enhanced susceptibility to harmful stimuli in aged tissue.

Senescence marker protein-30 knockout mouse as a novel murine model of senile lung.

Senescence marker protein-30 (SMP30) was originally identified as a novel protein of which expression decreases in an androgen-independent manner with aging in the rat liver and functions to protect cells from apoptosis. By reverse transcription-polymerase chain reaction analysis, SMP30 mRNA transcripts were found in the mouse lung, liver, kidney, testis and cerebrum. We examined SMP30 expression in the mouse liver, kidney and lung during aging and a distinct temporal profile of SMP30 expression was found in each tissue; the SMP30 mRNA level peaked at 1-3 months of age and decreased thereafter in the liver (the highest at 1 month of age followed by a rapid decline and consistently low thereafter in the kidney), and peaked at 12 months of age in the lung. To investigate the physiological role of SMP30 in the lung, immunohistochemical studies of wild-type (SMP30Y/+) mice and histopathological examinations of SMP30 knockout (SMP30Y/-) mice were performed. Immunoreactivity against anti-SMP30 antibody was mainly detected in bronchial epithelial cells and strongly detected at 6-12 months of age. Morphometric analysis was performed to measure the mean linear intercept and destructive index, and found peripheral airspace enlargement without alveolar destruction in SMP30Y/- mice at 1, 3 and 6 months of age compared with the SMP30Y/+ mice. Our results strongly suggest that SMP30Y/- mice could be a novel model for a senile lung and further examinations of SMP30Y/- mice may offer clues to elucidate the mechanisms of the development of pulmonary diseases in the elderly.

Senescence Marker Protein-30 Knockout Mouse Liver Is Highly Susceptible to Tumor Necrosis Factor-α- and Fas-Mediated Apoptosis

Senescence marker protein-30 (SMP30) is a calcium-binding protein that decreases in an androgen-independent manner with aging. To elucidate the physiological role of this protein, we introduced a null mutation of the SMP30 gene into the germ line of mice. Despite the complete lack of SMP30 (SMP30-/-), these mutant mice were indistinguishable from their wild-type (SMP30+/+) littermates in terms of development and fertilization capability. We then investigated the tissue susceptibility for apoptosis induced by cytokine using primary cultured hepatocytes, because SMP30 could rescue cells from cell death caused by calcium influx, using a calcium ionophore as previously described. SMP30-/- hepatocytes were found to be more susceptible to apoptosis induced by tumor necrosis factor-alpha (TNF-alpha) plus actinomycin D (ActD) than SMP30+/+ hepatocytes. In addition, the TNF-alpha/ActD-induced caspase-8 activity in SMP30-/- hepatocytes was twofold greater than that in SMP30+/+ hepatocytes. In contrast, no significant difference was observed in the TNF-alpha/ActD-induced nuclear factor-kappa B activation of SMP30+/+ versus SMP30-/- hepatocytes, indicating that SMP30 is not related to TNF-alpha/ActD-induced nuclear factor-kappa B activation itself. Moreover, deletion of the SMP30 gene enhanced liver injury after treatment in vivo with anti-Fas antibody and the SMP30+/- mice showed intermediate susceptibility to Fas-induced apoptosis. Collectively, these results demonstrate that SMP30 acts to protect cells from apoptosis.

Localization of senescence marker protein-30 (SMP30) in the mouse submandibular gland and histological findings in the SMP30 knock-out mouse

Senescence marker protein-30 (SMP30), which has a molecular mass of 30 kDa, is strongly expressed during maturation and decreases with aging in the liver and kidney. SMP30 is suspected of being closely related to cell functions, especially intracellular Ca2+ homeostasis, in differentiated tissues of the liver and kidney. We produced an antibody against SMP30 to study the localization of SMP30 in the submandibular gland of normal mice using an enzyme-labeled antibody. The submandibular gland was also observed under transmission electron microscopy in newly created SMP30-knockout mice compared to normal mice. SMP30 was found to be widely distributed in acinic cells of ductal and peripheral areas of the submandibular gland. Degenerative mitochondria tended to be abundantly observed in cells with SMP30, although marked changes were not noted in SMP30-knockout mice.