Gill microcirculation of the air-breathing climbing perch, Anabas testudineus (Bloch):relationships with the accessory respiratory organs and systemic circulation

The general macrocirculation and branchial microcirculation of the air-breathing climbing perch, Anabas testudineus, was examined by light and scanning electron microscopy of vascular corrosion replicas. The ventral aorta arises from the heart as a short vessel that immediately bifurcates into a dorsal and a ventral branch. The ventral branch distributes blood to gill arches 1 and 2, the dorsal branch to arches 3 and 4. The vascular organization of arches 1 and 2 is similar to that described for aquatic breathing teleosts. The respiratory lamellae are well developed but lack a continuous inner marginal channel. The filaments contain an extensive nutritive and interlamellar network; the latter traverses the filament between, but in register with, the inner lamellar margins. Numerous small, tortuous vessels arise from the efferent filamental and branchial arteries and anastomose with each other to form the nutrient supply for the filament, adductor muscles, and arch supportive tissues. The efferent branchial arteries of arches 1 and 2 supply the accessory air-breathing organs. Arches 3 and 4 are modified to serve primarily as large-bore shunts between the dorsal branch of the ventral aorta and the dorsal aorta. In many filaments from arches 3 and 4, the respiratory lamellae are condensed and have only 1-3 large channels. In some instances in arch 4, shunt vessels arise from the afferent branchial artery and connect directly with the efferent filamental artery. The filamental nutrient and interlamellar systems are poorly developed or absent. The respiratory and systemic pathways in Anabas are arranged in parallel. Blood flows from the ventral branch of the ventral aorta, through gill arches 1 and 2, into the accessory respiratory organs, and then returns to the heart. Blood, after entering the dorsal branch of the ventral aorta, passes through gill arches 3 and 4 and proceeds to the systemic circulation. This arrangement optimizes oxygen delivery to the tissues and minimizes intravascular pressure in the branchial and air-breathing organs. The efficiency of this system is limited by the mixing of respiratory and systemic venous blood at the heart.     

Succinic dehydrogenase activity of the respiratory muscles of a fresh-water teleost, Bagarius bagarius (Ham.) (Sisoridae, Pisces).

Functional morphology including the origin, insertion, and innervation of the respiratory muscles in relation to buccal pressure pump and opercular suction pumps in a fresh-water bottom dwelling siluroid fish, Bagarius bagarius have been studied. Histochemical studies were made on the succinic dehydrogenase activity of adductor mandibulae, retractor tentaculi, levator operculi, dilatator operculi, adductor operculi, intermandibularis, interhyoideus, hyohyoideus superior and constrictor branchialis. The intensity of reaction reveals the presence of three types of muscle fibres in some of the respiratory muscles. The muscle containing red muscle fibres are mostly innervated by the branches of the VIIth cranial nerve. The retractor tentaculi consists of superficial white muscle fibres and the interior part is dominated by red muscle fibres. The muscles (adductor operculi, levator operculi, dilatator operculi, interhyoideus, hyohyoideus superior) concerned with the opercular suction pumps are of mixed type and consist of white and red muscle fibres, whereas adductor mandibulae and intermandibularis are made up entirely of white muscle fibres. The adductor muscle bundles of the constrictor branchialis, which are responsible for movement of gill filaments, are dominated by the red muscle fibres. The abductor part, however, is made up entirely of white muscle fibres.     

A Three-dimensional Tissue Culture Model to Study Primary Human Bone Marrow and its Malignancies

Tissue culture has been an invaluable tool to study many aspects of cell function, from normal development to disease. Conventional cell culture methods rely on the ability of cells either to attach to a solid substratum of a tissue culture dish or to grow in suspension in liquid medium. Multiple immortal cell lines have been created and grown using such approaches, however, these methods frequently fail when primary cells need to be grown ex vivo. Such failure has been attributed to the absence of the appropriate extracellular matrix components of the tissue microenvironment from the standard systems where tissue culture plastic is used as a surface for cell growth. Extracellular matrix is an integral component of the tissue microenvironment and its presence is crucial for the maintenance of physiological functions such as cell polarization, survival, and proliferation. Here we present a 3-dimensional tissue culture method where primary bone marrow cells are grown in extracellular matrix formulated to recapitulate the microenvironment of the human bone (rBM system). Embedded in the extracellular matrix, cells are supplied with nutrients through the medium supplemented with human plasma, thus providing a comprehensive system where cell survival and proliferation can be sustained for up to 30 days while maintaining the cellular composition of the primary tissue. Using the rBM system we have successfully grown primary bone marrow cells from normal donors and patients with amyloidosis, and various hematological malignancies. The rBM system allows for direct, in-matrix real time visualization of the cell behavior and evaluation of preclinical efficacy of novel therapeutics. Moreover, cells can be isolated from the rBM and subsequently used for in vivo transplantation, cell sorting, flow cytometry, and nucleic acid and protein analysis. Taken together, the rBM method provides a reliable system for the growth of primary bone marrow cells under physiological conditions.     

Mutation of environmental mycobacteria to resist silver nanoparticles also confers resistance to a common antibiotic

Non-tuberculous mycobacteria are a threat to human health, gaining entry to the body through contaminated water systems, where they form persistent biofilms despite extensive attempts at disinfection. Silver is a natural antibacterial agent and in nanoparticle form activity is increased by a high surface area. Silver nanoparticles (AgNPs) have been used as alternative disinfectants in circulating water systems, washing machines and even clothing. However, nanoparticles, like any other antibiotic that has a pervasive durable presence, carry the risk of creating a resistant population. In this study Mycobacterium smegmatis strain mc²155 was cultured in AgNP enriched agar such that only a small population survived. Surviving cultures were isolated and re-exposed to AgNPs and AgNO₃ and resistance to silver was compared to a negative control. After only a single exposure, mutant M. smegmatis populations were resistant to AgNPs and AgNO₃. Further, the silver resistant mutants were exposed to antibiotics to determine if general resistance had been conferred. The minimum inhibitory concentration of isoniazid was four times higher for silver resistant mutants than for strain mc²155. However, core resistance was not conferred to other toxic metal ions. The mutants had lower resistance to CuSO₄ and ZnSO₄ than the mc²155 strain.     

Comparative studies of Ca2+-dependent proteases (CDP I and CDP II) from Allomyces arbuscula.

Allomyces arbuscula, an aquatic fungus, contains two Ca2+-dependent neutral cysteine proteases (CDP I and CDP II), eluting respectively, at 0.07 and 0.2 M NaCl from DEAE cellulose columns. The purified CDP I has a Mr of 39 kDa whereas CDP II appears as a doublet of 43 and 40 kDa. Both enzymes require free thiol, the same concentration of Ca2+ for half maximal activation, and are inactivated by thiol protease inhibitors. Our results show that despite these similarities the two enzymes are different because affinity-purified CDP II antibodies do not cross-react with CDP I antigen in Western blots. In contrast, there is a strong cross-reaction between the two 43 and 40 kDa CDP II peptides and their respective antibodies. Both enzymes cleave preferentially the carboxy terminus of Arg and to a limited extent Lys on the cleavage site. This primary specificity is governed by the nature of the amino acids in the P2 and P3 positions. In general either Pro or Gly in P2 is required, with preference for Pro and in P3 position, Gly over Val. CDP II has higher catalytic activity than CDP I. The sulfhydryl reagent NEM is a more potent inhibitor of CDP I than CDP II. Although the function of the phosphorylable site(s) is not clear, both CDP I and CDP II contain phosphorylable serine residue(s).     

Conformational flexibility and loss of structural rigidity for a model hexapeptide,GRGDTP: 1H‐NMR and molecular dynamics studies

The NMR and molecular dynamics methods are used to study the conformations of a hexapeptide, GRGDTP, which has been shown to be accessible to various types of cell‐adhesion based cellular behaviors such as cell‐to‐matrix interactions, cell differentiation, immunogenicity development, gene expression, angiogenesis, metastasis, sex determination and gamete fusion. ¹H‐NMR results indicate the existence of weak 5→2 hydrogen bonded β‐turn type‐III. Molecular simulation studies using a mixed protocol of distance geometry, constrained minimization, restrained molecular dynamics followed by energy minimization resulted additional conformations that include about 64% of population of inverse γ‐turn (HB, 3→1) and about 35% population of γ‐turn (HB, 4→2). The inter‐proton distances observed in γ‐and inverse γ‐turns are also consistent with the NMR constraints. The variable internal hydrogen bonding due to γ‐turns initiated at Gly 1 and Arg 2 , and its tendency to inter‐convert between γ‐and inverse γ‐turn conformations imply that the peptide is flexible in nature. © 2013 Wiley Periodicals, Inc. Biopolymers 99: 460–471, 2013.     

Thermostimulation of conidiation and succinic oxidative metabolism of Neurospora crassa

Summary The conidiogenic effect of temperature was investigated for its manifestation on the succinic oxidative system of Neurospora crassa. It was found that the initiation of conidiation was associated with a peak of succinate dehydrogenase activity. The succinate dehydrogenase activity decreased after the peak period of conidial differentiation. The strongest succinate dehydrogenase activity was measured in 37° C cultures.A second wave increase of succinate dehydrogenase was present in femalefertile (25° C) but was absent in the female-sterile (37° and 40° C) cultures.The cytochrome oxidase activity steadily decreased with the aging of the cultures.Malonate (10^-1 m) was found to stimulate conidiation. This stimulation was associated with higher succinate dehydrogenase activity.Riboflavine was found to accumulate with aging and was higher in 37° and 40° C cultures respectively as compared to 25° C cultures.The ultrathin sections of the conidia revealed that increase in conidial size at 37° C was accompanied with the swelling of the mitochondria and prominence of the endoplasmic reticulum. In conidia from 40° C cultures the mitochondria were shrunken and the endoplasmic reticulum broken.Dedicated with gratitude to Prof. Dr. A. Rippel-Baldes.