Regional Differences in Elements of Human Peroneus LongusTendons

Many studies have been performed on the structure, molecular composition, and biochemical properties of tendons. However, comparatively little research has been conducted on the content of various trace elements within tendons. Six elements were analyzed in four regions of the peroneus longus tendon: the tensional part of the tendon immediately proximal to the lateral malleolus (region A), the compressive region of the tendon in contact with the lateral malleolus (region B), the compressive region of the tendon in contact with the deep surface of the cuboid (region C), and the tensional part of the tendon between the cuboid and first metatarsal, to which the tendon is attached (region D). Regions B and C are wraparound regions. The calcium content was higher in region C (2.10?±?0.93 mg/g) than in both regions A (1.25?±?0.51 mg/g) and D (1.43?±?0.41 mg/g) (p?<?0.05), indicating that it is likely related to regional differences in cartilage degeneration. The phosphorus content was also higher in region C, possibly because of low alkaline phosphatase activity in this region. The sulfur content was higher in the wraparound regions (region B: 0.98?±?0.09 mg/g, region C: 1.24?±?0.19 mg/g) than in both regions A (0.83?±?0.11 mg/g) and D (0.83?±?0.1 mg/g) (p?<?0.01); sulfur content is thought to be influenced by tendon–bone compression. Finally, the magnesium content in the wraparound regions was also higher, which is probably related to a higher level of fibrocartilage. No significant relationships were found with regard to zinc or iron. Overall, the findings of the present study indicate that element contents are related to function and anatomical differences in tendons, and that they may even vary within the same tendon.     

Tuberous sclerosis complex 2 loss-of-function mutation regulates reactive oxygen species production through Rac1 activation

The products of the TSC1 (hamartin) and TCS2 (tuberin) tumor suppressor genes negatively regulate cell growth by inhibiting mTOR signaling. Recent research has led to the postulation that tuberin and/or hamartin are involved in tumor migration, presumably through Rho activation. Here we show that LEF-8 cells, which contain a Y1571 missense mutation in tuberin, express higher Rac1 activity than tuberin negative and positive cells. We also provide evidence of obvious lamellipodia formation in LEF-8 cells. Since the production of TSC2^Y1571H cannot form a hetero-complex with hamartin, we further analyzed another mutant, TSC2^R611Q, which also lacks the ability to form a complex with hamartin. Introducing both forms of mutated TSC2 into COS-1 cells increased Rac1 activity as well as cell motility. We also found these two mutants interacted with Rac1. We further demonstrated that the introduction of mutated TSC2 into COS-1 cells can generate higher reactive oxygen species (ROS). These results indicate that loss-of-function mutated tuberin can activate Rac1 and thereby increase ROS production.     

Identification of the sea urchin 350-kDa sperm-binding protein as a new sialic acid-binding lectin that belongs to the heat shock protein 110 family: implication of its binding to gangliosides in sperm lipid rafts in fertilization

The 350-kDa sperm-binding protein (SBP), a species-specific sperm-binding protein, is localized in the vitelline layer of sea urchin eggs. In this study, we have shown for the first time that sperm gangliosides are ligands for the intact glycosylated SBP. Using recombinant fragments of the SBP, the N-terminal heat shock protein 110-like domain was shown to be responsible for the binding. The intact SBP could bind various gangliosides, and the binding was sialidase-sensitive and inhibited by sialyllactose, thus indicating that it is the sialic acid-binding protein. Calcium and magnesium ions were not required but they did enhance the binding activity of SBP. The observation that bacterially expressed recombinant SBP and the sialidase-treated intact glycosylated SBP lost divalent cation-dependent enhancement of binding activity suggests that the sialylated carbohydrate moieties of the SBP may be involved in this property. Furthermore, the SBP was shown to bind sperm lipid rafts, in which gangliosides are enriched, and this binding was lost upon sialidase treatment of the lipid rafts. Finally, liposomes containing the ganglioside specifically inhibited fertilization. Taken together, these results allow us to identify SBP as a member of a new class of sialic acid-binding lectin belonging to the Hsp110 family, and indicate that SBP may be involved in interaction of sperm with the vitelline layer of the egg.     

Role of calnexin in the ER quality control and productive folding of CFTR; differential effect of calnexin knockout on wild-type and DeltaF508 CFTR

Cystic fibrosis (CF) is caused by the mutation in CF transmembrane conductance regulator (CFTR), a cAMP-dependent Cl(-) channel at the plasma membrane of epithelium. The most common mutant, DeltaF508 CFTR, has competent Cl(-) channel function, but fails to express at the plasma membrane since it is retained in the endoplasmic reticulum (ER) by the ER quality control system. Here, we show that calnexin (CNX) is not necessary for the ER retention of DeltaF508 CFTR. Our data show that CNX knockout (KO) does not affect the biosynthetic processing, cellular localization or the Cl(-) channel function of DeltaF508 CFTR. Importantly, cAMP-induced Cl(-) current in colonic epithelium from CNX KO/DeltaF508 CFTR mice was comparable with that of DeltaF508 CFTR mice, indicating that CNX KO failed to rescue the ER retention of DeltaF508 CFTR in vivo. Moreover, we show that CNX assures the efficient expression of WT CFTR, but not DeltaF508 CFTR, by inhibiting the proteasomal degradation, indicating that CNX might stimulate the productive folding of WT CFTR, but not DeltaF508 CFTR, which has folding defects.     

Effects of exposure to a circularly polarized 50-Hz magnetic field on plasma and pineal melatonin levels in rats

We sought to determine whether a 6-week exposure to a 50-Hz rotating magnetic field influences melatonin synthesis by 11–18 week-old Wistar-King male rats. Rats were exposed continuously to a rotating magnetic field at 1, 5, 50, or 250 μT (spatial vector rms) for 6 weeks, except for twice-weekly breaks of about 2 h for cleaning of cages and feeding. The rats were housed in exposure and sham-exposure facilities, which were located in the same room, under a 12:12 light-dark photoperiod (lights on at 06:00 h). The room was constantly illuminated by 4 small, dim red lights (< 0.07 lux in dark period). Levels of plasma and pineal gland melatonin were determined by radioimmunoassay. A significant decrease of melatonin was observed between the control group and groups exposed to a magnetic field at a flux density in excess of l μT during the night time, but no statistical differences were found among the exposed groups. These results indicate that subchronic exposure of albino rats to a 50-Hz rotating magnetic field influences melatonin production and secretion by the pineal gland. © 1993 Wiley-Liss, Inc.     

Enhancement of tumor cell susceptibility to lymphokine-activated killer cells by treatment with the streptococcal preparation OK432

We investigated whether tumor cell lysis by LAK cells was augmented by treatment with OK432in vitro. NK and LAK activity against K562 cells was not enhanced by their treatment with OK432. In contrast, the susceptibility of OK432-treated Daudi and KATO-III cells to lysis by LAK cells was enhanced. Succinate dehydrogenase activity and RNA synthesis were impaired in Daudi and KATO-III cells by treatment with OK432, and moreover the expression of HLA Class I antigen and ₂-microglobulin was inhibited in OK432-treated KATO-III cells. Thus, it is suggested that the enhancement of the susceptibility of OK432-treated tumor cells with regard to succinate dehydrogenase activity, RNA synthesis, and HLA Class I antigen expression.     

Neochilenin,a new glycoside of 3-O-methylquercetin,and other flavonols in the tepals ofNeochilenia,Neoporteria andParodia species (Cactaceae)

3-O-Methylated flavonols were isolated as crystals for the first time from the flowers ofNeochilenia, Neoporteria andParodia species belonging to the sub-family Cereoideae (Cactaceae), which are native to South America. The structures of three compounds were confirmed by chemical and spectral means. In the tepals of 7 species ofNeoporteria, 3-methyl ether of quercetin was found in the form of aglycone, whereas it was present as the 7-O-glucoside in the tepals ofParodia sanguiniflora and as the 4′-O-glucoside in the tepals of three species ofNeochilenia. Among those two glucosides of quercetin 3-methyl ether, the former has been found in a whole plant ofArtemisia transiliensis (Compositae), while the latter is new to the literature. Therefore, the term “neochilenin” may be assigned to this new pigment. Contribution from the Research Institute of Evolutionary Biology, No. 44.     

Drebrin is a novel connexin-43 binding partner that links gap junctions to the submembrane cytoskeleton

BACKGROUND: Connexins form gap junctions that mediate the transfer of ions, metabolites, and second messengers between contacting cells. Many aspects of connexin function, for example cellular transport, plaque assembly and stability, and channel conductivity, are finely tuned and likely involve proteins that bind to connexins' cytoplasmic domains. However, little is known about such regulatory proteins. To identify novel proteins that interact with the COOH-terminal domain of Connexin-43 (Cx43), the most widely expressed connexin family member, we applied a proteomics approach to screen fractions of mouse tissue homogenates for binding partners. RESULTS: Drebrin was recovered as a binding partner of the Cx43 COOH-terminal domain from mouse brain homogenate. Drebrin had previously been described as an actin binding protein that diminishes in brains during Alzheimer's disease. The novel Drebrin-Cx43 interaction identified by proteomics was confirmed by colocalization of endogenous proteins in astrocytes and Vero cells, coimmunoprecipitation, electron microscopy, electrophysiology, coexpression of both proteins with fluorescent tags, and live-cell FRET analysis. Depletion of Drebrin in cells with siRNA results in impaired cell-cell coupling, internalization of gap junctions, and targeting of Cx43 to a degradative pathway. CONCLUSIONS: We conclude that Drebrin is required for maintaining Cx43-containing gap junctions in their functional state at the plasma membrane. It is thus possible that Drebrin may interact with gap junctions in zones of cell-cell contacts in a regulated fashion in response to extracellular signals. The rearrangement or disruption of interactions between connexins and the Drebrin-containing submembrane cytoskeleton directs connexins to degradative cellular pathways.     

Accumulation of Inorganic Polyphosphate in phoU Mutants of Escherichia coli and Synechocystis sp. Strain PCC6803

The biological process for phosphate (P_i) removal is based on the use of bacteria capable of accumulating inorganic polyphosphate (polyP). We obtained Escherichia coli mutants which accumulate a large amount of polyP. The polyP accumulation in these mutants was ascribed to a mutation of the phoU gene that encodes a negative regulator of the P_i regulon. Insertional inactivation of the phoU gene also elevated the intracellular level of polyP in Synechocystis sp. strain PCC6803. The mutant could remove fourfold more P_i from the medium than the wild-type strain removed.