Characterization of alginate lyase (ALYII) from Pseudomonas sp. OS-ALG-9 expressed in recombinant Escherichia coli

An alginate lyase named ALYII was purified to homogeneity from Escherichia coli JM109 carrying a recombinant plasmid, pJK26 harbouring the alyII gene from Pseudomonas sp. OS-ALG-9 by column chromatography with DEAE-cellulose, CM-Sephadex C-50, butyl-Toyopearl 650 M and isoelectric focusing. The molecular size of the purified ALYII was estimated to be 79 kDa by SDS-PAGE and its pI was 8.3. The enzyme was most active at pH 7.0 and 30 °C. Its activity was completely inhibited by Hg²⁺. The enzyme was poly -D-1, 4-mannuronate-specific rather than -D-1, 4-guluronate-specific and it showed a promotion effect in alginate degradation by combination with ALY, an another poly -D-1, 4-mannuronate-specific alginate lyase from the same strain.     

Prolongation of survival of mice bearing the Eb and ESb lymphoma by treatment with interferon inducers alone or in combination with Corynebacterium parvum

Summary The objective of this study was to evaluate if pretreatment with Corynebacterium parvum (C. parvum) augments the effects of interferon (IFN) inducers on survival of DBA/2 mice transplanted with two syngeneic lymphoma variants, the low metastatic Eb and the high metastatic ESb tumor. The involvement of IFN in the treatment effects was investigated. As inducers of IFN-/ Newcastle disease virus (NDV), polyinosinic-polycytidylic acid (polyI:polyC), and 10-carboxymethyl-9-acridanone (CMA) were injected i. p. at the site of tumor transplantation. The Eb tumor was found to be sensitive to the antiproliferative action of IFN-/ in vitro. In vivo single injections of each of the inducers retarded growth of the Eb tumor. In C. parvum-pretreated mice the effects of the inducers on survival were markedly increased. There was a correlation between prolonged survival and local IFN levels in response to polyI:polyC or CMA but not upon NDV. Injections of each of the inducers increased cytotoxicity of peritoneal exudate cells against the Eb tumor cells in vitro especially when mice were pretreated with C. parvum. Although other mechanisms cannot be excluded IFN-mediated activation of host defence and also direct antiproliferative effects of endogenously produced IFN seem to be involved in the antitumor effects by these IFN inducers in the Eb model. In the ESb tumor model irrespective of additional pretreatment with C. parvum survival was only slightly prolonged by the treatments and endogenous IFN induction did not result in any real benefit for the animals. When compared with Eb cells the ESb cells were less sensitive to the antiproliferative action of IFN-/ in vitro and less sensitive to in vitro cytotoxicity by the host cells. Although other mechanisms may additionally be active in vivo the different susceptibility of the Eb and ESb tumor cells to the direct and indirect actions of IFN seems to contribute to the different responsiveness of these tumor cell lines to the treatments with IFN inducers.     

Lead exposure in Saudi Arabia and its relationship to smoking

Lead was determined in whole blood samples obtained from 202 Saudi male volunteers. The influence of smoking on lead exposure was investigated. Blood lead was significantly higher in current smokers than in non-smokers and previous smokers (P< 0.05). The distribution of blood lead data in the screened subjects suggested the eixstence of two mixed populations and a cut-off of 12 g dl ^–1 was found where the two populations separate. Of the exposed population, 80% with blood lead concentrations above 12 g dl^–1 were smokers.     

Structural Comparison of Human Mammalian Ste20-Like Kinases

Background

The serine/threonine mammalian Ste-20 like kinases (MSTs) are key regulators of apoptosis, cellular proliferation as well as polarization. Deregulation of MSTs has been associated with disease progression in prostate and colorectal cancer. The four human MSTs are regulated differently by C-terminal regions flanking the catalytic domains.

Principal Findings

We have determined the crystal structure of kinase domain of MST4 in complex with an ATP-mimetic inhibitor. This is the first structure of an inactive conformation of a member of the MST kinase family. Comparison with active structures of MST3 and MST1 revealed a dimeric association of MST4 suggesting an activation loop exchanged mechanism of MST4 auto-activation. Together with a homology model of MST2 we provide a comparative analysis of the kinase domains for all four members of the human MST family.

Significance

The comparative analysis identified new structural features in the MST ATP binding pocket and has also defined the mechanism for autophosphorylation. Both structural features may be further explored for inhibitors design.

Enhanced version

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Proteoglycan-type I collagen fibril interactions in bone and non-calcifying connective tissues

The association of proteogtycans with type I collagen fibrils in skin, tendon, cornea and bone has been determined by electron microscopy using an electrondense dye, Cupromeronic blue, in the critical electrolyte concentration mode, backed up by biochemical analysis and digestion by hyaluronidase or keratanase. A major proteoglycan of thesoft tissues, containing dermatan sulphat, is shown to be regularly and orthogonally arranged at the surface of the fibrils. Uranyl acetate counterstaining revealed that the main specific binding site is the d band, which previous work indicated is very close to the initial site of calcification of type I collagen fibrils. Bone, deminer-alized by a non-aqueous technique which preserves the proteoglycan in the tissue does not contain orthogonal arrays; the interfibrillar proteoglycan filaments are oriented parallel to the fibril axis. The main proteoglycan in bone is chondroitin sulphate-rich. It is suggested that dermatan sulphate proteoglycan plays a role in preventing soft connective tissues from calcifying.     

Multiple forms of inducible drug-metabolizing enzymes: A reasonable mechanism by which any organism can cope with adversity

Summary All organisms possess a number of genetically regulated mechanisms in order to cope with rapid adverse changes in the environment. The two systems which appear to respond to a seemingly endless array of chemical specificities are the immune response and the induction of drug-metabolizing enzymes. Similarities and differences between the immunoglobulin and the cytochrome P-450-mediated monooxygenase systems are described. DNA insertion sequences, plasmid transposons, maize controlling elements, gene duplication, intervening sequences, and high-frequency intergenic recombination are all discussed as possible methods by which organisms can adapt quickly to a new selective pressure. If the regulation of P-450 induction resembles in any way the other methods by which pro- and eukaryotes cope genetically with numerous forms of environmental adversity, therefore, it is very likely that mammalian tissues contain hundreds, if not thousands, of inducible forms of P-450.Portions of this overview were presented at the Symposium on The Effects of Drugs on Enzyme Induction Related to the Metabolism of Drugs and Carcinogenic Compounds, 14th Symposium Medicum Hoechst, County Mayo, Ireland, May, 1978 (1), and the Symposium on Isolated Drug-Metabolizing Enzymes, Mainz, Germany, July, 1978 (2). The author is Chief of the Developmental Pharmacology Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20205.     

Comparison of Immunity in Mice Cured of Primary/Metastatic Growth of EMT6 or 4THM Breast Cancer by Chemotherapy or Immunotherapy

Purpose

We have compared cure from local/metastatic tumor growth in BALB/c mice receiving EMT6 or the poorly immunogenic, highly metastatic 4THM, breast cancer cells following manipulation of immunosuppressive CD200:CD200R interactions or conventional chemotherapy.

Methods

We reported previously that EMT6 tumors are cured in CD200R1KO mice following surgical resection and immunization with irradiated EMT6 cells and CpG oligodeoxynucleotide (CpG), while wild-type (WT) animals developed pulmonary and liver metastases within 30 days of surgery. We report growth and metastasis of both EMT6 and a highly metastatic 4THM tumor in WT mice receiving iv infusions of Fab anti-CD200R1 along with CpG/tumor cell immunization. Metastasis was followed both macroscopically (lung/liver nodules) and microscopically by cloning tumor cells at limiting dilution in vitro from draining lymph nodes (DLN) harvested at surgery. We compared these results with local/metastatic tumor growth in mice receiving 4 courses of combination treatment with anti-VEGF and paclitaxel.

Results

In WT mice receiving Fab anti-CD200R, no tumor cells are detectable following immunotherapy, and CD4+ cells produced increased TNFα/IL-2/IFNγ on stimulation with EMT6 in vitro. No long-term cure was seen following surgery/immunotherapy of 4THM, with both microscopic (tumors in DLN at limiting dilution) and macroscopic metastases present within 14 d of surgery. Chemotherapy attenuated growth/metastases in 4THM tumor-bearers and produced a decline in lung/liver metastases, with no detectable DLN metastases in EMT6 tumor-bearing mice-these latter mice nevertheless showed no significantly increased cytokine production after restimulation with EMT6 in vitro. EMT6 mice receiving immunotherapy were resistant to subsequent re-challenge with EMT6 tumor cells, but not those receiving curative chemotherapy. Anti-CD4 treatment caused tumor recurrence after immunotherapy, but produced no apparent effect in either EMT6 or 4THM tumor bearers after chemotherapy treatment.

Conclusion

Immunotherapy, but not chemotherapy, enhances CD4⁺ immunity and affords long-term control of breast cancer growth and resistance to new tumor foci.     

Different effects of two brassinosteroids on growth,auxin and cytokinin content in tobacco callus tissue

Two synthetic brassinosteroids, 24-epibrassinolide (24-epiBR) and 2,3, 17-trihydroxy-5-androstan-6-one (THA-BR), exhibit different effects on growth of tobacco callus tissue. When added to a culture medium containing growth-limiting amounts of auxin, 24-epiBR reduced and THA-BR increased the fresh weight yield of tissue up to 53% and 207%, respectively, after 6 weeks of cultivation. The stimulatory and inhibitory effects of the two brassinosteroids on tissue growth occurred over a broad range of concentrations without a pronounced maximum corresponding to the yes or no type of response. Different effects of 24-epiBR and THA-BR on tissue growth were inversely proportional to the content of endogenous cytokinins. Maximum contents of predominant cytokinins N⁶-(²-isopentenyl)adenine (iP) and trans-zeatin (Z) in tissues supplied with 24-epiBR in growth-inhibiting concentrations were up to 3.7 fold and 3.4 fold higher, respectively, as compared to tissues grown on media containing growth-stimulating concentrations of THA-BR. Stimulation of tissue growth by THA-BR correlated with content of endogenous IAA and an inverse correlation was found between the content of endogenous IAA and cytokinins in tissues supplied with 24-epiBR. THA-BR exhibited weak cytokinin-like activity in a bioassay based on stimulation of growth of lateral buds of pea while 24-epiBR was inactive. Results indicate that the qualitatively different effects of the two brassinosteroids on growth of tobacco tissue may reflect their different influence on content of endogenous cytokinin.Abbreviations BR(s) brassinosteroid(s) - 24-epiBR 24-epibrassinolide - THA-BR 2,33, 17-trihydroxy-5-androstan-6-one - CK(s) cytokinin(s) - iP N⁶-(²-isopentenyl)adenine - [9R]iP N⁶-(²-isopentenyl)adenosine - Z trans-zeatin - [9R]Z ribosyl-trans-zeatin - ABA abscisic acid - IAA indole-3-acetic acid - NAA naphtalene-1-acetic acid - DEAE cellulose diethylaminoethyl cellulose - HPLC high performance liquid chromatography - ELISA enzyme linked immuno-sorbent assay     

Kinetic,ESR, and trapping evidence for in vivo binding of Mn(II) to glutamine synthetase in brain cells

Mn(II) has been proposed as a potential modulator of various important CNS enzymes, particularly glutamine synthetase, which is compartmentalized in the cytoplasm of glia. Previous studies demonstrated that total glial Mn(II) was 50–57 M, of which 30–40% occurs in the cytoplasm. In the present study, electron spin resonance (ESR) was used to determine that the concentration of free cytoplasmic Mn(II) in cultured chick glial cells is 0.8 (±0.2) M, very near K_d for the GS-Mn(II) complex. No free Mn(II) could be detected in glial mitochondria. Association of Mn(II) with brain glutamine synthetase (GS) was assessed under in vivo conditions in the presence of millimolar Mg(II) by trapping bound⁵⁴Mn(II) ions in the active site with irreversible inhibitors, namely methionine-sulfoximine (MSOX) or specific analogues thereof plus ATP. Ovine brain tissue was lysed directly into buffer containing Mn(II), 3 mM Mg(II), 1 mM MSOX, 1 mM ATP, 200 mM KCl, and 20 mM NaCl. Alternatively, primary cultures of chick glial cells were permeabilized into these inactivation mixtures. -Methyl-d,l-prothionine-S,R-sulfoximine was used to specifically inhibit the mechanistically-related enzyme -glutamyl-cysteine synthetase prior to specific inactivation of GS by -ethyl-d,l-methionine-S,R-sulfoximine. Even inthe presence of 2–3 mM Mg(II), with only 5–10 M Mn(II) present, approximately 20–30% of GS subunits were trapped with bound Mn(II). These results indicate that brain GS exhibits a high degree of specificity for binding Mn(II) over Mg(II) and that Mn(II) binds to GS to a significant extent under in vivo conditions.     

The protective role of DL-α-lipoic acid in the oxidative vulnerability triggered by Aβ-amyloid vaccination in mice

Recent reports indicate that -amyloid peptide (A) vaccine based therapy for Alzheimers disease (AD) may be on the horizon. There are however, concerns about the safety of this approach. Immunization with A has several disadvantages, because it crosses the blood brain barrier and cause inflammation and neurotoxicity. The present work is aimed to study the protective effective of -lipoic acid (LA) in the oxidative vulnerability of -amyloid in plasma, liver, spleen and brain, when A fibrils are given intraperitoneally in inflammation induced mice. Result shows that reactive oxygen species (ROS) in the astrocytes of inflammation induced mice along with A (IA) has shown 2.5-fold increase when compared with LA treated mice. The increased level of lipid peroxidase (LPO) (p < 0.05) and decreased antioxidant status (p < 0.05) were observed in the plasma, liver, spleen and brain of IA induced mice when compared with LA treated mice.Data shows that there were no significant changes observed between the control and LA treated mice. Our biochemical and histological results highlight that significant oxidative vulnerability was observed in IA treated mice, which was prevented by LA therapy. Our findings suggest that the antioxidant effect of LA when induced with A may serve as a potent therapeutic tool for inflammatory AD models. (Mol Cell Biochem 270: 29–37, 2005)     

Social and reproductive behavior of three mouthbrooding cardinalfishes,Apogon doederleini,A. niger and A. notatus

Synopsis Social and reproductive behavior of three paternal mouthbrooding cardinalfishes (Apogonidae) were investigated in the shallow marine waters of Shirahama, Japan. The solitary species Apogon doederleini and A. niger bred in transient pairs, in which a male and female associated for only a few hours of each afternoon on less than 5 successive days. The prespawning behavior was the same as the courtship display on days prior to spawning. After spawning, egg-incubating males were usually left alone. The gregarious species Apogon notatus formed territorial lasting pairs, which resided at given sites from dawn to dusk on each day during a period of a month or more. After spawning, the egg-incubating male either continued to stay with his mate in the territory, or left it to enter into an aggregation. In the latter case, the female continued to reside in the territory, pairing with a new male whom she brought from an aggregation. It is suggested that in paternal apogonids the prolonged pair bond and territoriality should have developed only in gregarious species as secondary adaptation for reproductive success: to avoid conspecific interference during spawning.     

Inheritance of heading date,plant height,ear length and spikelets per spike in an intervarietal cross of wheat

Summary A study to obtain information on early segregating generations of an intervarietal cross WG 357 X Tobari 66 in spring wheat on the genetics of days to heading, plant height, ear length and spikelets per spike was conducted. WG 357 has amber, hard and lustrous grains and is a well adapted high yielding variety of North India whereas Tobari 66 is red grained introduction from CIMMYT.The parental F₁, F₂, B₁, B₂, biparentals, F₃ (parents of biparentals), F₃ bulk and F₄ bulk generations were studied in order to provide analysis of generations means (Mather 1949; Hayman 1958) and variance component analysis (Kearsay 1965; Perkins and Jinks 1970).There were highly significant differences among the generations for all the characters studied. There were significant differences among the F₃ lines as well as among the biparental progenies. Only in case of ear length was the contrast between the two also significant. The mean value of most of the generations arising from the cross fell between the parental range.The three-parameter model failed to account for the variation in generation means in the case of days to heading. This character was concluded to be influenced by linkage and higher order interactions. For the other characters the three parameter model was adequate. For all characters, additive gene effects were most important as compared to dominance gene effects.The analysis of gene action as provided by the generation variance indicated that additive variance was much more pronounced as compared to dominance variance. The heritability was high for days to heading (71 per cent for narrow sense and 80 per cent in broad sense) and plant height (62 and 93 per cent in narrow and broad sense respectively.The implications of the results in breeding programmes have been discussed.     

Immunomodulatory effect of selenosemicarbazides and selenium inorganic compounds, distribution in organs after selenium supplementation

Antioxidant properties of selenium producing a protective barrier against free radicals play an important role in numerous metabolic and immunologic processes associated with oxidation- reduction reactions which take place during intracellular digestion of phagocyted bacteria. The aim of our study was to examine the properties of an organic compound of selenium, 4-(o-tolilo)- selenosemicarbazide of p-chlorobenzoic acid in terms of its retention in organs, effect on erythropoesis and phagocytic abilities of neutrophiles as well as antioxidant properties in neutrophiles tested with NBT test. This compound as well as inorganic sodium selenate was given to Swiss mice at the dose of 10^–3 g Se/kg for the period of 10 days. The concentrations of selenium in livers of mice treated with sodium selenate and selenosemicarbazide were found to be higher than in controls (18,7 g lg^–1 and 23.2 g lg^–1 vs. 12 g lg^–1, respectively). Analysis of blood cells count has shown a significant decrease in neutrophile levels in both groups treated with selenium. The influence of selenium compounds on phagocytosis and especially NBT test has been determined (3.8% of positive cells in the controls vs. 2.2% and 0.9% in the groups treated with sodium selenate and selenosemicarbazide, respectively). Our preliminary investigations suggest that selenosemicarbazides are biologically active compounds and can modify neutrophile functions.     

Activation of immunoglobulin control elements in trasgenic mice

To assess the role interleukins and mitogens play in regulating immunoglobulin (Ig) gene expression via the Ig enhancer and promoter, transgenic mice carrying two different Ig gene regulatory regions were generated. One, EkCAT, contains the Ig heavy chain enhancer (E) and the light chain promoter driving the chloramphenicol acetyltransferase (CAT) gene. In the other, EkCAT, CAT is under the control of the promoter alone. E and relative activity were assessed by CAT assay. In EkCAT mice, low CAT expression was consistently found in spleen, bone marrow, mesenteric lymph node, and thymus but not in brain, lung, or kidney. In EkCAT mice, CAT expression was detectable just above background in lymphoid tissues, suggesting a basic level of tissue specificity in the absence of the enhancer. Whole spleen cell cultures prepared from the mice were treated with lymphokines and mitogens. Lipopolysaccharide (LPS), concanavilin A (Con A), interleukin 6 (IL-6), and interferon- (IFN-) increased CAT expression to varying extents in cells derived from EkCAT mice but not in spleen cells prepared from EkCAT mice. Thus, the presence of E, in addition to the promoter, is essential for the stimulation of CAT expression mediated by these factors. B cells from EkCAT mice were separated by density into populations of small and large cells. In untreated small B cells, no CAT expression was detected and only addition of LPS resulted in an increase in CAT expression. In large B cells, CAT was expressed at a low level without addition of exogenous factors. Incubation with LPS, IL-6, Con A and IFN- caused CAT expression to increase several-fold. This transgenic system provides a means to identify exogenous factors that activate Ig enhancers and promoters.This work has been submitted in partial fulfillment of the requirements for the doctoral degree from the George Washington University.     

Induction of antifreeze protein production by juvenile hormone in larvae of the beetle,Dendroides canadensis

Summary Larvae of the beetleDendroides canadensis accumulate protein antifreezes during the winter.D. canadensis which were collected in the early fall, prior to the initiation of cold hardening processes, were treated with either 3.3 or 6.6 g juvenile hormone I topically in acetone and maintained for 21 days under normally non-inductive acclimation conditions (16 light/8 dark, 20 °C). Hormone treated animals significantly elevated the levels of antifreeze protein in their hemolymph compared to those of acetone treated and untreated controls or animals measured on the day of collection. D. canadensis treated with the anti-JH compound precocene II (P2) in acetone for 24 h at a concentration of 20 g/cm² (a dose below LD₅₀ for behavioral survival) and then maintained under acclimation conditions conducive to antifreeze protein production (8 light/16 dark, 20 °C) for 2 weeks failed to elevate levels of antifreeze. Acetone treated control animals accumulated a significant concentration of antifreeze protein.D. canadensis were also treated with 20 and 150 g/cm² P2 (a dose below the LD₅₀ for gross survival) followed by acclimation to short (8 h) photoperiod at 10 °C. All animals receiving the higher P2 dosage failed to elevate antifreezes while only 42.9% of the individuals treated with the lower dosage initiated antifreeze protein production. In contrast, over 80% of untreated and 70% of acetone treated controls responded to these inductive acclimation conditions by elevating antifreeze concentrations.These results indicate that juvenile hormone participates in the seasonal control of antifreeze protein production inDendroides canadensis. Since this species does not enter a diapause state prior to or throughout the winter this is the first evidence establishing a direct hormonal mechanism involved with insect cold hardiness.     

Stimulation of non-specific resistance to tumors in the mouse using a poly(maleic-acid-styrene)-conjugated neocarzinostatin

Summary The development of non-specific resistance to tumors following stimulation with poly(maleic-acid-styrene)-conjugated neocarzinostatin (SMANCS), a polymer-conjugated derivative of neocarzinostatin, was investigated in mice. The growth of syngeneic solid tumors (Meth-A fibrosarcoma and RL 1 leukemia) inoculated into BALB/c mice was suppressed after one treatment with SMANCS at doses ranging from 0.14 mg/kg to 3.4 mg/kg i.v. 24 h before tumor implantation. Since previously observations concerning SMANCS have shown that it disappeared within 1.5 h after i.v. administration in mice and that it was inactivated quickly in plasma, SMANCS evidently inhibited tumor growth by mediating non-specific resistance. In addition, the non-specific resistance to tumors stimulated by SMANCS could be passively transferred to untreated mice by serum which was shown to contain interferon (IFN) from 12 h to 20 h after SMANCS administration. However, the resistance was not produced by serum prepared from mice at 8 h or 32 h after administration presumably because of the observation that the interferon activity was only demonstrated from 12 h to 28 h after SMANCS stimulation. When the serum specimens were treated with anti-IFN- antiserum, the antitumor activity of the sera was abrogated. However, no significant change was detected in the antitumor activity of the specimens following treatment with anti-IFN-/ antiserum. Treatment of mice with SMANCS and anti-IFN- antiserum together resulted in the elimination of the non-specific resistance to tumors. The IFN induced in the sera of mice by SMANCS was shown to be 57% IFN- and 41% IFN-/. Half of the interferon produced in SMANCS-stimulated mice could be eliminated by treatment with anti-IFN-, and treatment of SMANCS-stimulated mice with both anti-IFN- and anti-IFN-/ antisera resulted in a total absence of detectable interferon. These findings suggest that while the administration of SMANCS induces both IFN- and IFN-/ production, in this case, it is only the former which mediates the non-specific resistance to tumors.     

Pharmacokinetics and in vitro and in vivo anti-tumor response of an interleukin-2-human serum albumin fusion protein in mice

Purpose: Albuleukin fusion protein is a recombinant human interleukin-2 (rIL-2) genetically fused to recombinant human serum albumin (rHSA). The pharmacokinetics and pharmacologic activity of Albuleukin were examined in mice to determine whether the fusion protein had the immunomodulatory and anti-tumor properties of rIL-2 as well as a prolonged serum half-life due to the rHSA. Methods: The effect of Albuleukin on lymphocyte proliferation, IL-2 receptor binding, and release of IFN- from human NK cells were examined in vitro. For the pharmacokinetic analysis, Albuleukin and rIL-2 were administered intravenously (i.v.) and subcutaneously (s.c.) to BALB/c mice, both at a single dose of 500 g/kg. The anti-tumor properties of Albuleukin were evaluated in a Renca tumor model in BALB/c mice and in a metastatic liver model of B16F10 melanoma in C57B1/6 mice. In the Renca tumor model, BALB/c mice were dosed intraperitoneally (i.p.) and s.c. with Albuleukin on days 12, 14, 16, 19, 21, and 23 and i.p. with rIL-2 daily for two periods of 5 days (days 10–14 and 17–21). In the B16 melanoma model, C57B1/6 mice were dosed s.c. with rIL-2 twice daily or Albuleukin every 48 h for 14 days. Results: In vitro, Albuleukin induced the proliferation of primary human and mouse T cells and B cells and primary human NK cells, competed with rIL-2 for binding to the IL-2 receptors, and induced the production of IFN- from primary human NK cells. The s.c. bioavailability of Albuleukin was about 45% relative to the i.v. dose. Plasma half-life was prolonged and ranged from 6 to 8 h with Albuleukin, compared to 19–57 min with rIL-2. Total clearance of Albuleukin was about 50-fold slower than that of rIL-2 after i.v. dosing. In vivo, Albuleukin suppressed the growth of Renca tumors and induced a dense infiltration of CD4⁺ and CD8⁺ T cells. Both Albuleukin and rIL-2 significantly reduced the tumor burden in mice with hepatic B16F10 metastases. Albuleukin significantly reduced the incidence of residual macroscopic hepatic tumors, resulting in improved survival relative to controls and rIL-2. Conclusion: Results from these studies suggest that the therapeutic efficacy of rIL-2 is improved in mice by prolonging its in vivo half-life through genetic fusion to albumin. Albuleukin, the fusion protein, had pronounced anti-tumor effects in Renca and hepatic melanoma tumor models without an increase in mortality. On the basis of its preclinical effects, Albuleukin was brought to the clinic to assess its therapeutic benefit in a variety of cancers.     

Single gene control of resistance to cutaneous leishmaniasis in mice

A series of inbred, congenic resistant, and hybrid strains of mice were intradermally inoculated with 10⁶ promastigotes of Leishmania tropica. These mice were divided into susceptible and resistant groups using the criteria of lesion size, development of metastatic foci and skin-test reactivity. At 16 weeks of infection, resistant strains A/J, DBA/1J, AKR/J, CBA/J, C3H/HeJ, NZB/BINJ, C57BL/6J, C57BL/10Sn, B10.D2, B10.129(10M), and B10.CE(30NX) had completely resolved their lesions, while susceptible SWR/J and BALB/cJ mice demonstrated large, nonhealing cutaneous lesions. In addition, BALB/cJ developed metastatic lesions on the extremities which progressively increased in size. All BALB/cJ and SWR/J mice died by 7 1/2 months of infection. The BALB/cJ x C57BL/6JF₁ hybrid behaved in an intermediate fashion showing a slower expansion of cutaneous ulcers and a delayed development of metastatic foci, however, the infection ultimately proved fatal. The F₂ generation could be separated into three distinct groups: resistant, intermediate, and susceptible mice with a lesion size distribution pattern in conformity with a 1:2:1 ratio. Male/female susceptibility differences were not noted. These data indicated that development of acquired resistance may be under the control of a single, autosomal gene. The gene did not appear to be H-2-, Ir-2-, or H-11-linked as is seen with Leishmania donovani infections.     

Gamma carbonic anhydrase like complex interact with plant mitochondrial complex I

We report the identification by two hybrid screens of two novel similar proteins, called Arabidopsis thaliana gamma carbonic anhydrase like1 and 2 (AtCAL1 and AtCAL2), that interact specifically with putative Arabidopsis thaliana gamma Carbonic Anhydrase (AtCA) proteins in plant mitochondria. The interaction region that was located in the N-terminal 150 amino acids of mature AtCA and AtCA like proteins represents a new interaction domain. In vitro experiments indicate that these proteins are imported into mitochondria and are associated with mitochondrial complex I as AtCAs. All plant species analyzed contain both AtCA and AtCAL sequences indicating that these genes were conserved throughout plant evolution. Structural modeling of AtCAL sequences show a deviation of functionally important active site residues with respect to CAs but could form active interfaces in the interaction with AtCAs. We postulate a CA complex tightly associated to plant mitochondrial complex.     

Variation in stable isotope signatures of seston and a zooplanktivorous fish in a eutrophic Chinese lake

Temporal and spatial changes in ¹³C and ¹⁵N of seston (mainly phytoplankton) and isotopic relationship between seston and the lake anchovy (Coilia ectenes) were studied in the large eutrophic freshwater Lake Chaohu in China. Much of the spatial and temporal variation in ¹³C of lake anchovies was explained by variation in seston, indicating a strong link between pelagic primary production and higher order consumers. Because the lake is shallow, there were no significant differences in ¹³C and ¹⁵N of seston between surface and overlying waters. Spatially, the relatively high ¹³C and ¹⁵N of seston in the western part of the lake might be due to high levels of anthropogenically derived N and C introduced from the surrounding cities through sewage drainage systems. The trophic position of the lake anchovy in the food web of Lake Chaohu was estimated to be 2.9–4.1 (3.5 ± 0.4), which agrees well with the previous stomach content analysis suggesting that the lake anchovy fed both on zooplankton and small planktivorous fishes.