Hepatocytes from newborn and weanling rats in monolayer culture: Isolation by perfusion,fibronectin-mediated adhesion,spreading, and functional activities

Summary The two-step collagenase perfusion method originally developed for the high yield isolation of parenchymal cells from adult rat livers has been adapted to rats of 1 day, 1 week, and 3 weeks of age. The use of this method to isolate hepatocytes from five or six rats of the respective ages demonstrated its reliability in terms of cell yield, percentage of single cells, and cell viability. In all cases, hepatocytes attach with high efficiency to fibronectin precoated dishes using serum-free culture medium. The dynamics of spreading is faster for newborn hepatocytes than adult ones. The functional integrity of these parenchymal liver cells was assessed by their capacity to secrete albumin and α-fetoprotein in serum-free medium and to express lactate dehydrogenase activity over a 24-hr period in primary culture. Part of this work was presented at the 30th Annual Meeting of the Tissue Culture Association, Seattle, June, 1979.     

Distribution and metabolism of adrenocorticotropin in the rat.

The time course of plasma bioactive adrenocorticotropin (ACTH) concentrations measured following two rapid injections of the hormone at doses of 7.5 and 22.5 mU/100 g, iv, and one infusion over a period of 80 min at a rate of 1.3 mU/min per 100 g, to male Sprague-Dawley rats whose endogenous release of ACTH had been blocked, leads to the conclusion that the hormone is distributed in two compartments. Indeed, the rapid fall of plasma ACTH concentrations in the early minutes following either the injections or the stop of the infusion is followed by a much slower phase. There is no significant difference between the measurements and the two-compartment model outputs. The model represents, on the average, the mean values of the measurements plus or minus 1 standard error for the single injections and plus or minus 1.2 standard error for the infusion.     

A new fluorometric method for determination of picomoles of inorganic phosphorus. Application to the renal tubular fluid

A new method for the determination of inorganic phosphorus (Pi) in 4 nl of renal tubular fluid is described. Phosphate is converted to hexadimolybdatophosphate, which is reacted with thiamine to produce a highly fluorescent thiochrome. The reaction is stable and reproducible. Problems of interference are minimal. Microdeterminations of Pi in the proximal tubules and glomerular filtrates of the rat yielded results similar to those published in the literature.     

Adrenocortical responses to ACTH (author's transl)]

To characterize by a mathematical model the dynamics of adrenocortical responses to ACTH, the concentration of plasma corticosterone was measured following a series of injections and perfusions of ACTH to dexamethasone-treated male rats. A similar slope was observed during the rise of plasma corticosterone following graded pulse inputs, and the steady-state corticosterone secretion rate was shown to increase proportionally to the perfusion rate of ACTH up to saturation corresponding to maximal secretion. The data will be used, concurrently with our model of the dynamics of plasma cortiscosterone, for the identification of a mathematical model of the adrenocortical response to ACTH.     

The effect of microbial mycolytic agents on Trichophyton rubrum

Chitinolytic microorganisms isolated from forest soil and from healthy gypsy moth larvae (Porthetria dispar (L.) were screened for their ability to lyse Trichophyton rubrum mycelia. A few of these isolates were mycolytic on both autoclaved and on actively growing, intact, T. rubrum mycelia. Supernatants from these isolates, utilizing live T. rubrum as the sole carbon source, showed the same mycolytic ability. Assays of the supernatants for enzymatic activity revealed exocellular, stable enzymes that releases reducing substances including N-acetylglucosamine from the mycelia.     

ADAM9 silencing inhibits breast tumor cell invasion in vitro

ADAM9 (A Disintegrin And Metalloproteinase 9) is a member of the ADAM protein family which contains a disintegrin domain. This protein family plays key roles in many physiological processes, including fertilization, migration, and cell survival. The ADAM proteins have also been implicated in various diseases, including cancer. Specifically, ADAM9 has been suggested to be involved in metastasis. To address this question, we generated ADAM9 knockdown clones of MDA-MB-231 breast tumor cells using silencing RNAs that were tested for cell adhesion, proliferation, migration and invasion assays. In RNAi-mediated ADAM9 silenced MDA-MB-231 cells, the expression of ADAM9 was lower from the third to the sixth day after silencing and inhibited tumor cell invasion in matrigel by approximately 72% when compared to control cells, without affecting cell adhesion, proliferation or migration. In conclusion, the generation of MDA-MB-231 knockdown clones lacking ADAM9 expression inhibited tumor cell invasion in vitro, suggesting that ADAM9 is an important molecule in the processes of invasion and metastasis.     

Computational Simulations of the Early Steps of Protein Aggregation

There is strong evidence that the oligomers of key proteins, formed during the early steps of aggregation, could be the primary toxic species associated with human neurodegenerative diseases, such as Alzheimer''s and prion diseases. Here, we review recent progress in the development of computational approaches in order to understand the structures, dynamics and free energy surfaces of oligomers. We also discuss possible research directions for the coming years.Key Words: protein aggregation, simulations, amyloid fibril, oligomers, coarse-grained model, inhibitors     

Ultrashort nanosecond electric pulses evoke heterogeneous patterns of Ca2+ release from the endoplasmic reticulum of adrenal chromaffin cells

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Isolation and 16S rRNA sequence analysis of the beneficial bacteria from the rhizosphere of rice

The present study deals with the isolation of plant growth promoting rhizobacteria (PGPR) from rice (variety NIAB IRRI-9) and the beneficial effects of these inoculants on two Basmati rice varieties. Nitrogen-fixing activity (acetylene-reduction activity) was detected in the roots and submerged shoots of field-grown rice variety NIAB IRRI-9. Estimation of the population size of diazotrophic bacteria by ARA-based MPN (acetylene reduction assay-based most probable number) in roots and shoots indicated about 10(5)-10(6) counts/g dry weight at panicle initiation and grain filling stages. Four bacterial isolates from rice roots and shoots were obtained in pure culture which produced phytohormone indoleacetic acid (IAA) in the growth medium. Among these, three isolates S1, S4, and R3 reduced acetylene to ethylene in nitrogen-free semi-solid medium. Morphological and physiological characteristics of the isolates indicated that three nitrogen-fixing isolates S1, S4, and R3 belonged to the genus Enterobacter, while the non-fixing isolate R8 belonged to the genus Aeromonas. 16S rRNA sequence of one isolate from root (R8) and one isolate from shoot (S1) was obtained which confirmed identification of the isolates as Aeromonas veronii and Enterobacter cloacae, respectively. The 1517-nucleotide-long sequence of the isolate R8 showed 99% similarity with Aeromonas veronii (accession No. AF099023) while partial 16S rRNA sequence (two stretches of total 1271 nucleotide length) of S1 showed 97% similarity with the sequence of Enterobacter cloacae (accession No. AJ251469). The seedlings of two rice varieties Basmati 385 and Super Basmati were inoculated with the four bacterial isolates from rice and one Azospirillum brasilense strain Wb3, which was isolated from wheat. In the rice variety Basmati 385, maximum increase in root area and plant biomass was obtained in plants inoculated with Enterobacter S1 and Azospirillum Wb3, whereas in the rice variety Super Basmati, inoculation with Enterobacter R3 resulted in maximum increase of root area and plant biomass. Nitrogen fixation was quantified by using 15N isotopic dilution method. Maximum fixation was observed in Basmati 385 with the inoculants Azospirillum Wb3 and Enterobacter S1 where nearly 46% and 41% of the nitrogen was derived from atmosphere (%Ndfa), respectively. In general, higher nitrogen fixation was observed in variety Basmati 385 than in Super Basmati, and different bacterial strains were found more effective as inoculants for the rice varieties Basmati 385 and Super Basmati.