Functional and phenotypic analyses of interleukin 2-activated tumor-infiltrating lymphocytes

The tumor-infiltrating lymphocytes (TILs) were cultured with interleukin 2 (IL-2) to induce the activated killer cells possessing autologous tumor-killing activity, and analysed their cell surface phenotypes and assessed anti-tumor killing activity. Furthermore, the activated TILs were transferred into 7 patients adoptively resulting in complete remission in a patient with pancreatic cancer and partial remission in another patient with gastric cancer.The cytotoxic activities of activated TILs at 3 weeks-incubation was 72 ± 15, 42 ± 26, 27 ± 21 and 25 ± 15% against K562, Daudi, KATO-III and autologous tumor, respectively. The negative selection method, indicated that the killer cells recognizing autologous tumor cells consisted of CD4- or CD8-positive T lymphocytes and CD16- or CD56-positive natural killer cells. The activated TILs could not only lyse cultured tumor cell lines, but also autologous tumor cells.     

TENA, a selective kappa opioid receptor antagonist

A number of opioid antagonists (TENA, naloxone, Mr 2266, WIN 44441) were evaluated for their selectivity in antagonizing the effect of mu, kappa, and delta agonists in the guinea pig ileum (GPI) and mouse vas deferens (MVD) preparations. Among these four antagonists, TENA was the most potent and the only ligand which was selective for kappa receptors. In this regard TENA was approximately 27-times more effective in antagonizing the kappa agonist, U-50488H, relative to the mu agonist, morphine, and it was about 5-times more effective against ethylketazocine (EK) relative to morphine. At the same concentration (20 nM) TENA did not significantly antagonize the delta agonist, [D-Ala2,D-Ala5]enkephalin (DADLE), in the MVD. Also, TENA was more effective than naloxone, EK, or U-50488H in protecting kappa receptors from irreversible blockage by beta-CNA. The results of this study indicate that TENA is the most selective kappa antagonist yet reported.     

Analysis of feeding mechanism in a pitcher ofNepenthes hybrida

The process of digestion of captured feeds in a pitcher, an insect-trapping organ, ofNepenthes was studied. Changes in bacterial population, pH and NH₄ ⁺ concentrations in pitcher juice were examined. Strong activities of both acid- and alkaline phosphatase, phosphoamidase, esterase C4 and esterase C8 were found in the pitcher juice. Optimum pH of proteases in the juice and those secreted from bacteria showed pH 3.0 and pH 8.0–9.0, respectively. Twenty six strains of bacteria were isolated from 4 pitchers: 10 strains were gram positive, 16 strains were gram negative (10 strains had casein hydrolase activity). A proton excretion was induced by NH₄ ⁺ released from the added solutions, and accordingly, the pH of the solutions fell. As a simulation model of the digestion process of feeds in pitcher juice and polypeptone solution was added into the washed pitcher. A good correlation was found among the NH₄ ⁺ concentration, pH and bacterial cell titer.     

Localization of the antigen for the monoclonal antibody ②9F11-B-E4 interspecific cross-reaction in the freshwater triclads

The localization of the antigen for monoclonal antibody 9F11-B-E₄ was clarified by immuno-electron microscopy. The antigens were localized on the mitochondria and Golgi bodies in the male germ cells and on the secretory granules of various glands cells in the penis bulb and subepidermal parenchymal tissue of Phagocata vivida. The results of the interspecific cross-reaction tests with seven other freshwater triclads showed that these secretory granules are species-specific. A positive interspecific reaction was showed with Dugesia (family Dugesiidae), but not with Polycelis within the same family Planariidae which suggests the position of Phagocata within the Planariidae needs to be reassesed.     

Modulation of acute morphine tolerance by corticotropin-releasing factor and dynorphin A in the mouse spinal cord.

Previously, we have demonstrated that intrathecally (i.t.) administered corticotropin-releasing factor (CRF) in mice produces stimulus-specific antinociception and modulation of morphine-induced antinociception by mechanisms involving spinal kappa opioid receptors. Recently, we also have found that CRF releases immunoreactive dynorphin A, a putative endogenous kappa opioid receptor agonist, from superfused mice spinal cords in vitro. Dynorphin A administered intracerebroventricularlly (i.c.v.) to mice has been shown to modulate the expression of morphine tolerance. In the present study, the possible modulatory effects of i.t. administered CRF as well as dynorphin A on morphine tolerance were studied in an acute tolerance model. Subcutaneous administration of 100 mg/kg of morphine sulfate (MS) to mice caused an acute tolerance to morphine-induced antinociception. The antinociceptive ED50 of MS was increased from 4.4 mg/kg (naive mice) to 17.9 mg/kg (4 hours after the injection of 100 mg/kg MS). To study the modulatory effects of spinally administered CRF and dynorphin A on the expression of morphine tolerance, CRF and dynorphin A were injected i.t. at 15 min and 5 min, respectively, before testing the tolerant mice by the tail-flick assay. The antinociceptive ED50 of MS in tolerant mice was decreased to 8.8 mg/kg and 7.1 mg/kg, respectively, after i.t. administration of CRF (0.1 nmol) and dynorphin A (0.2 nmol). In contrast, 0.5 nmol of alpha-helical CRF (9-41), a CRF antagonist and 0.4 nmol of norbinaltorphimine, a highly selective kappa opioid receptor antagonist, when administered i.t. at 15 min before the tail-flick test in tolerant mice, increased the antinociceptive ED50 of MS to 56.6 mg/kg and 88.8 mg/kg, respectively. These data confirmed the modulatory effect of dynorphin A on morphine tolerance and suggested that CRF, which releases dynorphin A in several central nervous system regions, also plays a modulatory role in the expression of morphine tolerance.     

Protective mechanism of sodium molybdate against the acute toxicity of cadmium in rats. II. Prevention of cytoplasmic acidification

In order to clarify the protective mechanism of sodium molybdate against the acute toxicity of cadmium chloride in rat, the effect of in vivo sodium molybdate pretreatment on the cytotoxic action of cadmium in isolated hepatocytes was studied. The cytosolic pH of hepatocytes isolated from untreated rats immediately decreased with incubation in either neutral Hank's balanced salt solution (HBS), pH 7.4, containing 5 µM cadmium chloride minimum or acidic HBS (pH 7.1, 6.8, 6.5, and 6.2). The presence of 5 µM cadmium in HBS adjusted to pH 7.1 aggravated cytosalic acidification induced by the acidic medium alone. Cell viability of hepatocytes incubated in HBS at pH 6.2 was significantly reduced as compared to that of control cells in HBS at pH 7.4, but the presence of cadmium in the acidic HBS had no aggravating action against such a toxic action of the acidic medium although cellular uptake of the metal in the medium increased, as compared to that in HBS at pH 7.4. Molybdenum pretreatment alleviated cytoplasmic acidification induced by the treatment with HBS at pH 7.4 or 7.1 containing cadmium or by extracellular acid load wothout cadmium. This pretreatment also prevented the loss of cell viability induced by the treatment with HBS at pH 6.2 but could not attenuate that when cadmium was present in the medium.These facts suggest that molybdenum pretreatment alleviated the acute toxicity of cadmium in rat by preventing cytoplasmic acidification caused by the harmful metal.     

Isolation of selective 3H-chlornaltrexamine-bound complexes,possible opioid receptor components in brains of mice

The nonequilibrium narcotic antagonist, chlornaltrexamine (CNA) was used to bind selectively and covalently pioid specific sites on brain membrane preparations. Selective binding of [³H]CNA occured with a saturation maximum of 185 fmol/mg protein. Bound [³H]CNA was extracted with Triton X-100, dialyzed against Brij 36T, precipitated with trichloroacetic acid and chromatographed on an ultrogel AcA 22 column. The elution profile suggests that this extract contains a minimum of four selective [³H]CNA complexes. At least two of these complexes migrate in a single large peak. Column calibration showed that this peak eluted at 590,000 daltons. One of these specific [³H]CNA complexes elutes at the elution volume of the column and is dialyzable. Finally, putative aggregate of these complexes elutes with the void volume.     

Scanning Electron Microscopy of Rhizobium trifolii Infection Sites on Root Hairs of White Clover

White clover root hairs which were inoculated with Rhizobium trifolii 4S (infectious strain) contained infection threads which were observed by light microscopy and scanning electron microscopy. Three morphological types of root hairs retaining infection threads were recognized. The bacteria were strongly attached between the surfaces of two plant cell walls as follows: between surfaces of a root hair tip curled back on itself, between a protuberance from a root hair and its cell surface, or between two root hair tips clinging together. An anatomical analysis documented the attachment site of the infection thread sheath from the inside of the root hair cell.