Purpose: Telomere length (TL) is an indicator of age;however, hormonal influences complicate individual aging. It remains unclear whether TL shortening is a direct factor in both individual and cellular aging. Therefo...Purpose: Telomere length (TL) is an indicator of age;however, hormonal influences complicate individual aging. It remains unclear whether TL shortening is a direct factor in both individual and cellular aging. Therefore, we examined the direct relationship between TL and cellular senescence at the cellular level. Methods: Telomerase activity, TL, and gene expression were measured in cultured human lung-, fetal-, and skin-derived fibroblasts, human skin keratinocytes, and telomerase reverse transcriptase (TERT) gene-immortalized cells using detection kits, Cawthon’s method, and reverse transcription-quantitative polymerase chain reaction, respectively. Novel substances that elongate telomeres were screened to confirm cell rejuvenation effects. Results: Long-term cell culture of TIG-1-20 normal human fibroblasts resulted in TL shortening, decreased division rate, and senescence progression, whereas in OUMS-36T-2 cells, TL elongation via TERT gene transfer increased the division rate, reduced endoplasmic reticulum stress, and upregulated genes associated with young individuals, indicating that cellular rejuvenation occurs via TL elongation. In addition, a honey child powder (HCP) extract was found through screening, and the HCP extract strongly suppressed the menin gene, resulting in increased telomerase activity and extended cell lifespan. Upon addition of the HCP extract to skin fibroblasts, gene expression of moisturizing components, including collagen, hyaluronic acid, and elastin, increased, and exhibited a rejuvenating effect with an increase in elastin amount. Conclusions: TL elongation or shortening is involved in cell proliferation rate and cellular aging, and TL elongation rejuvenates cells. In addition, HCP extract has a rejuvenating effect on cells and is expected to be a rejuvenating compound.展开更多
Beige adipocytes are believed to have a high ability to consume fat. As such, compounds capable of inducing the development of beige adipocytes may be useful as drugs for anti-obesity and anti-type 2 diabetes. However...Beige adipocytes are believed to have a high ability to consume fat. As such, compounds capable of inducing the development of beige adipocytes may be useful as drugs for anti-obesity and anti-type 2 diabetes. However, the true nature of beige adipocytes remains unclear. The purpose of this study is to confirm whether or not white adipocyte can differentiate to beige adipocytes and to clarify the characteristics of beige adipocytes. We first searched for an inducer of beige adipocytes and found that kikyo extract, a component of bofu-tsusho-san, was a strong inducer. We then attempted to prove that beige adipocytes could be induced from white adipocytes. Second, we clarified the characteristics of beige adipocytes induced from white adipocytes. The results suggested that beige adipocytes were high-performance adipocytes with a greater ability to synthesize and consume triglyceride and take up glucose than white adipocytes.展开更多
Obese subjects show both a fatty predisposition and a higher risk of metabolic syndrome. This characteristic depends on adipocytes. However, the roles of adipocytes in metabolic syndrome have been insufficiently inves...Obese subjects show both a fatty predisposition and a higher risk of metabolic syndrome. This characteristic depends on adipocytes. However, the roles of adipocytes in metabolic syndrome have been insufficiently investigated, because few pure adipocyte cell lines have been isolated. The present study had two objectives: the isolation of a pure adipocyte cell line and clarification of the differences between adipocytes and preadipocytes, and screening for compounds that can potentially prevent metabolic syndrome. We isolated a novel adipocyte cell line, 3T3-L24. In the 3T3-L24 cells, the gene expression levels (of C/EBPα and β, PPARγ and AP2) and the production of triglyceride and TNFα were much higher than those in the preadipocyte 3T3-L1 cells. We used the 3T3-L24 adipocytes to screen for compounds that could inhibit triglyceride accumulation and TNFα secretion. Fatty acids enhanced the triglyceride accumulation. Sodium carboxylate, taurine and carnitine not only inhibited triglyceride accumulation, but also inhibited TNFα secretion. Therefore, these compounds might be effective to decrease the risk of metabolic syndrome in obese subjects.展开更多
Cancer tissues contain cancer stem cells (CSCs), which play important roles in cancer metastasis. However, the mechanisms through which cancer cells dedifferentiate into stem cells have not yet been elucidated. In thi...Cancer tissues contain cancer stem cells (CSCs), which play important roles in cancer metastasis. However, the mechanisms through which cancer cells dedifferentiate into stem cells have not yet been elucidated. In this study, the effects of high concentrations of polyamines produced in cancer cells on dedifferentiation were examined. The results showed that when normal human fibroblasts were cultured with high concentrations of spermine, the obtained polyamine-induced cells expressed alkaline phosphatase and marker proteins of pluripotent stem cells, although apoptosis occurred in most cells. In contrast, another polyamine-induced stem (PIS) cell line (Spe-2 PIS cells), obtained by culture in medium containing Rock, p53, and Bax inhibitors plus spermine, did not show signs of apoptosis. These Spe-2 PIS cells expressed marker proteins of pluripotent stem cells and differentiated into cardiomyocytes, brown adipocytes, and nerve cells. These results suggest that a high concentration of spermine, which often induces apoptosis in normal cells, has the capacity to dedifferentiate somatic cells into pluripotent stem cells and may be associated with the dedifferentiation of cancer cells, which continuously produce high concentrations of spermine. Moreover, the procedure to obtain Spe-2 PIS cells, which is simple and efficient, may have potential applications in regenerative medicine.展开更多
Vascular calcification, which causes occlusion and rupture of the vascular, is often observed in patients in the advanced stages of arteriosclerosis. One of the best procedures for inhibiting the accumulation of vascu...Vascular calcification, which causes occlusion and rupture of the vascular, is often observed in patients in the advanced stages of arteriosclerosis. One of the best procedures for inhibiting the accumulation of vascular calcification is to obstruct the differentiation of mesenchymal stem cells (MSCs) and/or vascular endothelial cells (VECs) in the vascular to osteoblasts. In this study, we evaluated the biochemical and genetic characteristics of the process of differentiation of MSCs and VECs to osteoblasts. C3H10T1/2 MSCs, TKD2 VECs and MC3T3-E1 preosteoblasts (POBs) were cultured in medium containing both hydrocortisone and glycerophosphate. These compounds showed strong effects promoting the differentiation of VECs as well as POBs, although the effect was weak in the MSCs. Moreover, C3H10T1/2 MSCs and TKD2 VECs were cultured in medium containing 10 mM retinol, after which the alkali phosphatase (ALP) activity of the MSCs and production of calcified nodules of TKD2 were significantly increased, whereas the marker genes for the osteoblasts were not. These results suggest that retinol does not have an effect in inducing the differentiation of VECs to osteoblasts, but rather exhibits a strong promoting effect on differentiation.展开更多
Obese individuals exhibit much higher risks not only for metabolic syndrome, but also for cancer and allergies, than normal-weight subjects. This fact suggests that signals secreted from adipocytes change the characte...Obese individuals exhibit much higher risks not only for metabolic syndrome, but also for cancer and allergies, than normal-weight subjects. This fact suggests that signals secreted from adipocytes change the characteristics of lymphocytes, such as macrophages and T-cells. We focused on a free fatty acid, oleic acid, as a signal inducing such changes and examined its effects on murine J774.2 macrophages. When the cells were cultured in medium containing high concentrations (1, 2 and 4 mM) of oleic acid, apoptosis occurred, and the apoptotic cells were gathered into clusters of very large size by the work of enzymes for phagocytosis. When the cells were cultured in medium containing 0.5 mM of oleic acid, the fatty acid did not affect cell growth;however, it inhibited nitrogen monoxide (NO) secretion and the gene expressions of interleukins and TNF-α. NO disturbs the invasion of macrophages into blood vessels, and interleukins promote the differentiation and proliferation of T- and B-cells. Therefore, these results suggest that the high risks for cancer and allergies observed in obese subjects are associated with the dysfunction of macrophages induced by fatty acids. Moreover, we also examined the protective effects of carnitine against dysfunction. However, carnitine did not exhibit sufficient effects.展开更多
Although Yarrowia lipolytica is an important host strain, there have so far been few studies on the production of glutathione by the strain. We therefore performed a study to obtain an improved strain of Y. lipolytica...Although Yarrowia lipolytica is an important host strain, there have so far been few studies on the production of glutathione by the strain. We therefore performed a study to obtain an improved strain of Y. lipolytica ATCC20688, which could produce a high yield of glutathione. First, the capability of glutathione production in the ATCC20688 strain was estimated. In comparison with other yeasts, the yield of this strain was higher than those in Pichia strains. Furthermore, this strain could produce glutathione by assimilating sodium oleate. We next performed mutation and gene cloning to improve the yield. After the yield of glutathione was improved in the isolated methylglyoxal-resistant mutant (MGR3), the glutathione synthetase gene was cloned into the MGR3 strain. By using this recombinant strain, we could reach the maximum yield and intracellular content of glutathione of 54 mg/L-medium and 30 mg/g-dry cell weight, respectively.展开更多
We searched for a superior melamine-degrading bacterium for the bioremediation of melamine. Cyanuric acid, which is a by-product produced during the biodegradation of melamine, shows strong nephrotoxicity. Therefore, ...We searched for a superior melamine-degrading bacterium for the bioremediation of melamine. Cyanuric acid, which is a by-product produced during the biodegradation of melamine, shows strong nephrotoxicity. Therefore, the melamine-degrading bacterium is also required to show a high ability to degrade cyanuric acid. We selected a melamine-degrading strain (MEL1) among ten cyanuric acid-degrading bacteria isolated from the soil. The species of MEL1 strain was Microbacterium esteramaticum or was extremely similar to that species, and the enzymatic activity of the melamine deaminase in the MEL1 strain was similar to that in the NRRLB-12227 strain. The ability of the MEL1 strain to degrade cyanuric acid was higher than its ability to degrade melamine, and therefore, the accumulation of by-products (ammeline, ammelide and cyanuric acid) during the degradation of melamine was minimal. These results suggest that the MEL1 strain is useful for the bioremediation of melamine.展开更多
文摘Purpose: Telomere length (TL) is an indicator of age;however, hormonal influences complicate individual aging. It remains unclear whether TL shortening is a direct factor in both individual and cellular aging. Therefore, we examined the direct relationship between TL and cellular senescence at the cellular level. Methods: Telomerase activity, TL, and gene expression were measured in cultured human lung-, fetal-, and skin-derived fibroblasts, human skin keratinocytes, and telomerase reverse transcriptase (TERT) gene-immortalized cells using detection kits, Cawthon’s method, and reverse transcription-quantitative polymerase chain reaction, respectively. Novel substances that elongate telomeres were screened to confirm cell rejuvenation effects. Results: Long-term cell culture of TIG-1-20 normal human fibroblasts resulted in TL shortening, decreased division rate, and senescence progression, whereas in OUMS-36T-2 cells, TL elongation via TERT gene transfer increased the division rate, reduced endoplasmic reticulum stress, and upregulated genes associated with young individuals, indicating that cellular rejuvenation occurs via TL elongation. In addition, a honey child powder (HCP) extract was found through screening, and the HCP extract strongly suppressed the menin gene, resulting in increased telomerase activity and extended cell lifespan. Upon addition of the HCP extract to skin fibroblasts, gene expression of moisturizing components, including collagen, hyaluronic acid, and elastin, increased, and exhibited a rejuvenating effect with an increase in elastin amount. Conclusions: TL elongation or shortening is involved in cell proliferation rate and cellular aging, and TL elongation rejuvenates cells. In addition, HCP extract has a rejuvenating effect on cells and is expected to be a rejuvenating compound.
文摘Beige adipocytes are believed to have a high ability to consume fat. As such, compounds capable of inducing the development of beige adipocytes may be useful as drugs for anti-obesity and anti-type 2 diabetes. However, the true nature of beige adipocytes remains unclear. The purpose of this study is to confirm whether or not white adipocyte can differentiate to beige adipocytes and to clarify the characteristics of beige adipocytes. We first searched for an inducer of beige adipocytes and found that kikyo extract, a component of bofu-tsusho-san, was a strong inducer. We then attempted to prove that beige adipocytes could be induced from white adipocytes. Second, we clarified the characteristics of beige adipocytes induced from white adipocytes. The results suggested that beige adipocytes were high-performance adipocytes with a greater ability to synthesize and consume triglyceride and take up glucose than white adipocytes.
文摘Obese subjects show both a fatty predisposition and a higher risk of metabolic syndrome. This characteristic depends on adipocytes. However, the roles of adipocytes in metabolic syndrome have been insufficiently investigated, because few pure adipocyte cell lines have been isolated. The present study had two objectives: the isolation of a pure adipocyte cell line and clarification of the differences between adipocytes and preadipocytes, and screening for compounds that can potentially prevent metabolic syndrome. We isolated a novel adipocyte cell line, 3T3-L24. In the 3T3-L24 cells, the gene expression levels (of C/EBPα and β, PPARγ and AP2) and the production of triglyceride and TNFα were much higher than those in the preadipocyte 3T3-L1 cells. We used the 3T3-L24 adipocytes to screen for compounds that could inhibit triglyceride accumulation and TNFα secretion. Fatty acids enhanced the triglyceride accumulation. Sodium carboxylate, taurine and carnitine not only inhibited triglyceride accumulation, but also inhibited TNFα secretion. Therefore, these compounds might be effective to decrease the risk of metabolic syndrome in obese subjects.
文摘Cancer tissues contain cancer stem cells (CSCs), which play important roles in cancer metastasis. However, the mechanisms through which cancer cells dedifferentiate into stem cells have not yet been elucidated. In this study, the effects of high concentrations of polyamines produced in cancer cells on dedifferentiation were examined. The results showed that when normal human fibroblasts were cultured with high concentrations of spermine, the obtained polyamine-induced cells expressed alkaline phosphatase and marker proteins of pluripotent stem cells, although apoptosis occurred in most cells. In contrast, another polyamine-induced stem (PIS) cell line (Spe-2 PIS cells), obtained by culture in medium containing Rock, p53, and Bax inhibitors plus spermine, did not show signs of apoptosis. These Spe-2 PIS cells expressed marker proteins of pluripotent stem cells and differentiated into cardiomyocytes, brown adipocytes, and nerve cells. These results suggest that a high concentration of spermine, which often induces apoptosis in normal cells, has the capacity to dedifferentiate somatic cells into pluripotent stem cells and may be associated with the dedifferentiation of cancer cells, which continuously produce high concentrations of spermine. Moreover, the procedure to obtain Spe-2 PIS cells, which is simple and efficient, may have potential applications in regenerative medicine.
文摘Vascular calcification, which causes occlusion and rupture of the vascular, is often observed in patients in the advanced stages of arteriosclerosis. One of the best procedures for inhibiting the accumulation of vascular calcification is to obstruct the differentiation of mesenchymal stem cells (MSCs) and/or vascular endothelial cells (VECs) in the vascular to osteoblasts. In this study, we evaluated the biochemical and genetic characteristics of the process of differentiation of MSCs and VECs to osteoblasts. C3H10T1/2 MSCs, TKD2 VECs and MC3T3-E1 preosteoblasts (POBs) were cultured in medium containing both hydrocortisone and glycerophosphate. These compounds showed strong effects promoting the differentiation of VECs as well as POBs, although the effect was weak in the MSCs. Moreover, C3H10T1/2 MSCs and TKD2 VECs were cultured in medium containing 10 mM retinol, after which the alkali phosphatase (ALP) activity of the MSCs and production of calcified nodules of TKD2 were significantly increased, whereas the marker genes for the osteoblasts were not. These results suggest that retinol does not have an effect in inducing the differentiation of VECs to osteoblasts, but rather exhibits a strong promoting effect on differentiation.
文摘Obese individuals exhibit much higher risks not only for metabolic syndrome, but also for cancer and allergies, than normal-weight subjects. This fact suggests that signals secreted from adipocytes change the characteristics of lymphocytes, such as macrophages and T-cells. We focused on a free fatty acid, oleic acid, as a signal inducing such changes and examined its effects on murine J774.2 macrophages. When the cells were cultured in medium containing high concentrations (1, 2 and 4 mM) of oleic acid, apoptosis occurred, and the apoptotic cells were gathered into clusters of very large size by the work of enzymes for phagocytosis. When the cells were cultured in medium containing 0.5 mM of oleic acid, the fatty acid did not affect cell growth;however, it inhibited nitrogen monoxide (NO) secretion and the gene expressions of interleukins and TNF-α. NO disturbs the invasion of macrophages into blood vessels, and interleukins promote the differentiation and proliferation of T- and B-cells. Therefore, these results suggest that the high risks for cancer and allergies observed in obese subjects are associated with the dysfunction of macrophages induced by fatty acids. Moreover, we also examined the protective effects of carnitine against dysfunction. However, carnitine did not exhibit sufficient effects.
文摘Although Yarrowia lipolytica is an important host strain, there have so far been few studies on the production of glutathione by the strain. We therefore performed a study to obtain an improved strain of Y. lipolytica ATCC20688, which could produce a high yield of glutathione. First, the capability of glutathione production in the ATCC20688 strain was estimated. In comparison with other yeasts, the yield of this strain was higher than those in Pichia strains. Furthermore, this strain could produce glutathione by assimilating sodium oleate. We next performed mutation and gene cloning to improve the yield. After the yield of glutathione was improved in the isolated methylglyoxal-resistant mutant (MGR3), the glutathione synthetase gene was cloned into the MGR3 strain. By using this recombinant strain, we could reach the maximum yield and intracellular content of glutathione of 54 mg/L-medium and 30 mg/g-dry cell weight, respectively.
文摘We searched for a superior melamine-degrading bacterium for the bioremediation of melamine. Cyanuric acid, which is a by-product produced during the biodegradation of melamine, shows strong nephrotoxicity. Therefore, the melamine-degrading bacterium is also required to show a high ability to degrade cyanuric acid. We selected a melamine-degrading strain (MEL1) among ten cyanuric acid-degrading bacteria isolated from the soil. The species of MEL1 strain was Microbacterium esteramaticum or was extremely similar to that species, and the enzymatic activity of the melamine deaminase in the MEL1 strain was similar to that in the NRRLB-12227 strain. The ability of the MEL1 strain to degrade cyanuric acid was higher than its ability to degrade melamine, and therefore, the accumulation of by-products (ammeline, ammelide and cyanuric acid) during the degradation of melamine was minimal. These results suggest that the MEL1 strain is useful for the bioremediation of melamine.
