Possible implication of metalloproteinases in post-mortem tenderization of fish muscle

ABSTRACT: It is suspected that the proteolytic breakdown of extracellular matrix proteins is responsible for the postmortem tenderization of fish muscle during chilled storage. In order to identify the type(s) of proteinases involved in this phenomenon, the effect of proteinase inhibitors, EDTA (ethylenediamine tetraacetic acid), 1,10-phenanthroline, ρ-APMSF [(ρ-amidinophenyl) methanesulfonyl fluoride hydrochloride] and E-64 [ L - trans -epoxysuccinyl-leucylamido-(4-guanidinobutane)] on tenderization was investigated by using Japanese flounder. Proteinase inhibitor solution was injected into a blood vessel in a caudal portion of live flounder and the firmness of muscle was then evaluated as a shear force value at 0 h and 6 h after death. Metalloproteinase inhibitors, EDTA and 1,10-phenanthroline, significantly suppressed postmortem tenderization. These findings suggest that metalloproteinases are candidates for proteinases involved in the postmortem tenderization of fish muscles. Although not significantly, p -APMSF, a serine proteinase inhibitor, partially suppressed muscle tenderization, which suggests that serine proteinases are also implicated in postmortem tenderization. A cysteine proteinase inhibitor, E-64, showed no effect, suggesting that cysteine proteinases are not involved.     

Endoscopic findings on alimentary lymphoma in 7 dogs

Alimentary lymphoma was evaluated endoscopically in 7 dogs and a histopathological examination was made to detect the origin of neoplastic lymphocytes in 5 dogs. A solitary mass in the rectum (1 case), irregular cobblestone appearance in the duodenum (4 cases) and a moderate irregular appearance resembling lymphocytic-plasmacytic enteritis (2 cases) were endoscopically detected. Endoscopic ultrasonography demonstrated increased thickness of the duodenal wall in 2 cases examined. Neoplastic lymphocytes of alimentary lymphoma proved to originate in T cells in all 5 cases examined by immunohistochemical analysis.     

Contemporary Strategies for the Synthesis of Tetrahydropyran Derivatives: Application to Total Synthesis of Neopeltolide,a Marine Macrolide Natural Product

Tetrahydropyrans are structural motifs that are abundantly present in a range of biologically important marine natural products. As such, significant efforts have been paid to the development of efficient and versatile methods for the synthesis of tetrahydropyran derivatives. Neopeltolide, a potent antiproliferative marine natural product, has been an attractive target compound for synthetic chemists because of its complex structure comprised of a 14-membered macrolactone embedded with a tetrahydropyran ring, and twenty total and formal syntheses of this natural product have been reported so far. This review summarizes the total and formal syntheses of neopeltolide and its analogues, highlighting the synthetic strategies exploited for constructing the tetrahydropyran ring.     

Animal cellulases with a focus on aquatic invertebrates

Cellulose is utilized as a nutritional source by various organisms. For a long time it was believed that only protozoa, bacteria, and fungi, in addition to plants and photosynthetic bacteria, are able to synthesize cellulases encoded by their own genes. However, the widespread distribution of cellulases throughout the animal kingdom has recently been recognized. Conventionally, animals digest cellulose utilizing cellulases derived from symbiotic bacteria in the digestive organs. However, recent molecular biological studies have shown that some cellulase genes are actually encoded on animal chromosomes. In addition, the homologous primary structure of cellulases obtained from various invertebrate phyla indicates the possible vertical transfer of the cellulase gene from ancient organisms that are now extinct. The results of studies on cellulases with unique enzymatic properties are expected to be applied to bioethanol production and aquaculture. In the present review, we describe cellulases, focusing primarily on aquatic invertebrates in which both endogenous and exogenous cellulases are involved in the breakdown of cellulose in the digestive organs.     

Therapeutic efficacy of ABT-737, a Bcl-2 inhibitor, in a canine melanoma cell line

The small molecule inhibitor, ABT-737, inhibits Bcl-2 that is overexpressed in many tumor cell lines and, in combination with an anticancer drug, can strongly enhance proapoptotic activity. In the present study, we evaluated the inhibitory activity of ABT-737 on the survival of a canine melanoma cell line (MCM-N1). MCM-N1 cell viability was decreased following 24- and 48-hr culture with ABT-737, depending on ABT-737 concentration, while cell viability was unchanged in controls. ABT-737 synergized with carboplatin to promote cell death. Notably, approximately 50% of MCM-N1 cells survived following culture with 2-4 μg/ml of carboplatin; whereas, less than 20% of MCM-N1 cells survived following culture with ABT-737 (1 mM) plus carboplatin (2-10 μg/ml).     

Comparison of the functions of the barley nicotianamine synthase gene HvNAS1 and rice nicotianamine synthase gene OsNAS1 promoters in response to iron deficiency in transgenic tobacco

Barley ( Hordeum vulgare L.) nicotianamine synthase gene ( HvNAS1 ) expression in barley is strongly induced by Fe deficiency in the roots and rice ( Oryza sativa L.) nicotianamine synthase gene ( OsNAS1 ) expression in rice is induced by Fe deficiency both in the roots and in the shoots. In dicots, NAS genes are not strongly induced by Fe deficiency, and they function to maintain Fe homeostasis. Rice OsNAS1promoter::GUS or barley HvNAS1promoter::GUS was introduced into tobacco ( Nicotiana tabacum L.) and tissue specificities and systemic regulation of their expression were compared. A split-root experiment revealed that the HvNAS1 promoter exhibited functions similar to those of Fe-acquisition-related genes in tobacco roots, suggesting that this promoter responded to certain Fe-deficiency systemic signals and to the Fe concentration in the rhizosphere. The HvNAS1 promoter might harbor a type of universal system of gene expression for Fe acquisition. However, the OsNAS1 promoter did not respond to local application of Fe to the roots and induced GUS activities in mature leaves in response to Fe deficiency. This promoter might possess numerous types of cis -acting sequences that are involved in Fe metabolism.     

Groundwater Deterioration Caused by Induced Recharge: Field Survey and Verification of the Deterioration Mechanism by Stochastic Numerical Simulation

Our field survey showed that thequality of shallow groundwater around the KatsuraRiver in the Kyoto Basin was strongly affected by theinfiltration of river water. Furthermore, that thedeterioration of the groundwater in the southern areato the west of the Katsura River may be related to theincrease in groundwater extraction. To clarify themechanism of groundwater deterioration, we havedeveloped a stochastic method to simulate groundwaterflow. The results showed that there was a largereduction in the groundwater level where groundwaterextraction was intense and recharge flowed from theKatsura River to the high extraction areas in thesouthern region. Another simulation showed that if thegroundwater extraction was 10% of the present removalrate, there would be little recharge from the KatsuraRiver into the groundwater and the quality of thegroundwater would be improved. Thus, we conclude thatthe cause of groundwater deterioration is probably dueto the induced recharge of deteriorated river waterfrom the Katsura River.     

The effect of small interfering RNA (siRNA) against the Bcl-2 gene on apoptosis and chemosensitivity in a canine mammary gland tumor cell line

The aim of this study was to investigate whether downregulation of Bcl-2 expression by small interfering RNA (siRNA) against the canine Bcl-2 gene would enhance the apoptosis and sensitivity of a canine mammary gland tumor cell line (CF33) to doxorubicin. Transfections of CF33 with siRNA were performed using cationic liposomes. Sequence-specific downregulation of Bcl-2 expression was measured by semiquantitative RT-PCR and Western blot analysis. Total viable cells were determined by MTS assay and apoptotic cell rates were determined by the immunohistochemical analysis on ssDNA. Our data showed the siRNA downregulated Bcl-2 expression which increased apotosis and also increased the sensitivity of CF33 to doxorubicin. This study indicated that downregulation of Bcl-2 expression by siRNA would be useful as a new protocol to increase the effect of doxorubicin on treatment of canine mammary gland tumors, requiring a detailed evaluation of siRNA in vivo.     

Construction of artificial promoters highly responsive to iron deficiency

Iron deficiency-responsive element 1 (IDE1) and IDE2 are cis-acting elements that are responsible for Fe-deficiency-inducible and root-specific expression of the barley (Hordeum vulgare L.) gene IDS2 (Fe-deficiency-specific clone no. 2). Using these cis-acting elements, we aimed to construct super-promoters that would induce prominent gene expression in the roots of Fe-deficient rice plants (Oryza sativa L.). Modules containing IDE1 and IDE2 of the IDS2 promoter were used as repeats or were linked to the Fe-deficiency-responsive promoter of barley IDS3, and were connected to known enhancer-like sequences. Five artificial promoters, as well as the native promoters of barley IDS2 or IDS3, were connected individually upstream of β-glucuronidase (GUS) and were introduced into rice. Transgenic rice plants were grown under control or Fe-deficient conditions, and GUS expression was analyzed. The artificial promoter that contained one module of IDE1 and IDE2 conferred strong Fe-deficiency-inducible GUS expression to the roots of rice plants. Each of the five artificial promoters induced a similar level of GUS expression in Fe-deficient roots, which did not exceed the GUS expression driven by the native IDS2 or IDS3 promoter. Artificial and native promoters induced GUS expression in response to Fe-deficiency in leaves, although the level of expression was lower than that in roots. Histochemical observations revealed that GUS expression driven by artificial and native promoters was spatially similar, and expression was dominant within vascular bundles and root exodermis. These findings suggest that there is coordinated expression of the genes that are involved in Fe-deficiency-induced Fe uptake in rice.