We studied the effects of various 111In-water soluble chelates and incubation media on labeling e... more We studied the effects of various 111In-water soluble chelates and incubation media on labeling efficiency of platelets and in vitro platelet aggregability. High labeling efficiency of platelets in ACD-saline was achieved with 111In-oxine sulfate, 111In-tropolone and 111In-MPO (2-mercaptopyridine-N-oxide). In the condition with 4.8 x 10(6)/mm3 platelets in ACD-plasma, 111In-oxine-sulfate had low labeling efficiency and inconsistent labeling, while 111In-tropolone and 111In-MPO had high labeling efficiency. In vitro platelet aggregability (ADP 2 microM) was reduced when platelets were labeled in the absence of plasma. However, there was no significant difference in platelet aggregability among 111In-platelets labeled by three different chelates. In conclusion, to maintain aggregation activity of the platelets with relatively high labeling efficiency, the best result was obtained by using MPO or tropolone chelate in plasma at 4.8 x 10(6)/microliters platelet concentration.
Following intraperitoneal injection of [3H]proline and colchicine, rat liver cells were dispersed... more Following intraperitoneal injection of [3H]proline and colchicine, rat liver cells were dispersed by collagenase perfusion and fractionated by low-speed and density gradient centrifugation. Analysis of the collagenous components using purified bacterial collagenase showed that 0.1 to 0.2% of the labeled protein produced by the liver cells was collagen. Considering the population of the hepatocytes in the liver (70%), 80% of the collagen produced by the liver could be attributed to the hepatocytes.
Type specific rabbit antibodies to bovine type I, II, III, and IV (basement membrane) collagens s... more Type specific rabbit antibodies to bovine type I, II, III, and IV (basement membrane) collagens showing no cross-reaction with other types of collagen were prepared by cross-adsorption and diethylaminoethyl-cellulose chromatography. The antibodies to bovine type I and III collagens showed a high cross-reaction with the corresponding human collagens, but those to type II and IV collagens did moderate and no cross-reactions with human type II and IV collagens, respectively. By using these antibodies, tissue distribution of various types of collagen in normal bovine lung was examined by indirect immunofluorescence microscopy. both type I and III collagens were found to distribute widely in the interstitium of bronchial tree, bronchial lamina propria and of interlobules as well as alveolar nipples and adventitia of pulmonary arteries. Type II collagen was located only in bronchial cartilage. The tissues mainly stained for type III collagen were the alveolar interstitium (also stained faintly for type I collagen) and the intima and media of the arteries. Type IV collagen was located in a membranous fashion in alveolar septa and bronchial smooth muscles and subepithelial layers as well as capillaries and the intima and media of arteries.
We reported previously that the forced expression of the chemokine BRAK, also called CXCL14 in he... more We reported previously that the forced expression of the chemokine BRAK, also called CXCL14 in head and neck squamous cell carcinoma (HNSCC) cells decreased the rate of tumor formation and size of tumor xenografts compared with mock-vector treated cells in athymic nude mice or in severe combined immunodeficiency mice. This suppression occurred even though the growth rates of these cells were the same under in vitro culture conditions, suggesting that a high expression level of the gene in tumor cells is important for the suppression of tumor establishment in vivo. The aim of this study was to determine whether CXCL14/BRAK transgenic mice show resistance to tumor cell xenografts or not. CXCL14/BRAK cDNA was introduced into male C57BL/6 J pronuclei, and 10 founder transgenic mice (Tg) were obtained. Two lines of mice expressed over 10 times higher CXCL14/BRAK protein levels (14 and 11 ng/ml plasma, respectively) than normal blood level (0.9 ng/ml plasma), without apparent abnormality. The sizes of Lewis lung carcinoma and B16 melanoma cell xenografts in Tg mice were significantly smaller than those in control wild-type mice, indicating that CXCL14/BRAK, first found as a suppressor of tumor progression of HNSCC, also suppresses the progression of a carcinoma of other tissue origin. Immunohistochemical studies showed that invasion of blood vessels into tumors was suppressed in tumor xenografts of CXCL14/BRAK Tg mice. These results indicate that CXCL14/BRAK suppressed tumor cell xenografts by functioning paracrine or endocrine fashion and that CXCL14/BRAK is a very promising molecular target for tumor suppression without side effects.
Morphogenesis is an old, and one of the latest, fascinating fields in biological science and a hu... more Morphogenesis is an old, and one of the latest, fascinating fields in biological science and a huge number of papers on molecular mechanisms underlying it have been published. But most of the works and reviews on these mechanisms pertain to molecules of, as it were, the planning or design of morphogenesis, such as morphogens and homeodomain proteins. In this review, I will describe the function of extracellular matrix (ECM) and other cell adhesion molecules in morphogenesis as that of actual morpho‐creating molecules, morphocreators, and discuss their roles as positional information‐pertaining molecules.
A two-dimensional mapping method developed for collagen chains has been applied to the CNBr-pepti... more A two-dimensional mapping method developed for collagen chains has been applied to the CNBr-peptides of rat collagen types I, II, and III and bovine type V (AB) collagen. The method consists of isoelectric focusing in a 4% polyacrylamide gel-6 M urea-2% Pharmalyte with a pH gradient ranging 5 to 10 in the first dimension and SDS-polyacrylamide slab gel (15%) electrophoresis at pH 8.8 in the second dimension. Characteristic peptide maps were obtained for each type of alpha-chain and can be used for identification of isolated collagen chains. Application of this method to an authentic mixture of collagen types and a fraction isolated from rat skin has indicated that the method is sensitive enough to resolve collagen types with a sample as low as 40 micrograms per alpha-chain.
We studied the effects of various 111In-water soluble chelates and incubation media on labeling e... more We studied the effects of various 111In-water soluble chelates and incubation media on labeling efficiency of platelets and in vitro platelet aggregability. High labeling efficiency of platelets in ACD-saline was achieved with 111In-oxine sulfate, 111In-tropolone and 111In-MPO (2-mercaptopyridine-N-oxide). In the condition with 4.8 x 10(6)/mm3 platelets in ACD-plasma, 111In-oxine-sulfate had low labeling efficiency and inconsistent labeling, while 111In-tropolone and 111In-MPO had high labeling efficiency. In vitro platelet aggregability (ADP 2 microM) was reduced when platelets were labeled in the absence of plasma. However, there was no significant difference in platelet aggregability among 111In-platelets labeled by three different chelates. In conclusion, to maintain aggregation activity of the platelets with relatively high labeling efficiency, the best result was obtained by using MPO or tropolone chelate in plasma at 4.8 x 10(6)/microliters platelet concentration.
Following intraperitoneal injection of [3H]proline and colchicine, rat liver cells were dispersed... more Following intraperitoneal injection of [3H]proline and colchicine, rat liver cells were dispersed by collagenase perfusion and fractionated by low-speed and density gradient centrifugation. Analysis of the collagenous components using purified bacterial collagenase showed that 0.1 to 0.2% of the labeled protein produced by the liver cells was collagen. Considering the population of the hepatocytes in the liver (70%), 80% of the collagen produced by the liver could be attributed to the hepatocytes.
Type specific rabbit antibodies to bovine type I, II, III, and IV (basement membrane) collagens s... more Type specific rabbit antibodies to bovine type I, II, III, and IV (basement membrane) collagens showing no cross-reaction with other types of collagen were prepared by cross-adsorption and diethylaminoethyl-cellulose chromatography. The antibodies to bovine type I and III collagens showed a high cross-reaction with the corresponding human collagens, but those to type II and IV collagens did moderate and no cross-reactions with human type II and IV collagens, respectively. By using these antibodies, tissue distribution of various types of collagen in normal bovine lung was examined by indirect immunofluorescence microscopy. both type I and III collagens were found to distribute widely in the interstitium of bronchial tree, bronchial lamina propria and of interlobules as well as alveolar nipples and adventitia of pulmonary arteries. Type II collagen was located only in bronchial cartilage. The tissues mainly stained for type III collagen were the alveolar interstitium (also stained faintly for type I collagen) and the intima and media of the arteries. Type IV collagen was located in a membranous fashion in alveolar septa and bronchial smooth muscles and subepithelial layers as well as capillaries and the intima and media of arteries.
We reported previously that the forced expression of the chemokine BRAK, also called CXCL14 in he... more We reported previously that the forced expression of the chemokine BRAK, also called CXCL14 in head and neck squamous cell carcinoma (HNSCC) cells decreased the rate of tumor formation and size of tumor xenografts compared with mock-vector treated cells in athymic nude mice or in severe combined immunodeficiency mice. This suppression occurred even though the growth rates of these cells were the same under in vitro culture conditions, suggesting that a high expression level of the gene in tumor cells is important for the suppression of tumor establishment in vivo. The aim of this study was to determine whether CXCL14/BRAK transgenic mice show resistance to tumor cell xenografts or not. CXCL14/BRAK cDNA was introduced into male C57BL/6 J pronuclei, and 10 founder transgenic mice (Tg) were obtained. Two lines of mice expressed over 10 times higher CXCL14/BRAK protein levels (14 and 11 ng/ml plasma, respectively) than normal blood level (0.9 ng/ml plasma), without apparent abnormality. The sizes of Lewis lung carcinoma and B16 melanoma cell xenografts in Tg mice were significantly smaller than those in control wild-type mice, indicating that CXCL14/BRAK, first found as a suppressor of tumor progression of HNSCC, also suppresses the progression of a carcinoma of other tissue origin. Immunohistochemical studies showed that invasion of blood vessels into tumors was suppressed in tumor xenografts of CXCL14/BRAK Tg mice. These results indicate that CXCL14/BRAK suppressed tumor cell xenografts by functioning paracrine or endocrine fashion and that CXCL14/BRAK is a very promising molecular target for tumor suppression without side effects.
Morphogenesis is an old, and one of the latest, fascinating fields in biological science and a hu... more Morphogenesis is an old, and one of the latest, fascinating fields in biological science and a huge number of papers on molecular mechanisms underlying it have been published. But most of the works and reviews on these mechanisms pertain to molecules of, as it were, the planning or design of morphogenesis, such as morphogens and homeodomain proteins. In this review, I will describe the function of extracellular matrix (ECM) and other cell adhesion molecules in morphogenesis as that of actual morpho‐creating molecules, morphocreators, and discuss their roles as positional information‐pertaining molecules.
A two-dimensional mapping method developed for collagen chains has been applied to the CNBr-pepti... more A two-dimensional mapping method developed for collagen chains has been applied to the CNBr-peptides of rat collagen types I, II, and III and bovine type V (AB) collagen. The method consists of isoelectric focusing in a 4% polyacrylamide gel-6 M urea-2% Pharmalyte with a pH gradient ranging 5 to 10 in the first dimension and SDS-polyacrylamide slab gel (15%) electrophoresis at pH 8.8 in the second dimension. Characteristic peptide maps were obtained for each type of alpha-chain and can be used for identification of isolated collagen chains. Application of this method to an authentic mixture of collagen types and a fraction isolated from rat skin has indicated that the method is sensitive enough to resolve collagen types with a sample as low as 40 micrograms per alpha-chain.
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