Rat lymphoma cell bioassay for prolactin: Observations on its use and comparison with radioimmunoassay

Lawson, David M.; Sensui, Naoto; Haisenleder, D.H.; Gala, Richard R.

doi:10.1016/0024-3205(82)90076-5

Cited by 50 publications

(19 citation statements)

References 7 publications

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“…FCS. On the following day, the cells were pre-treated for 30 min with rhPRL (200 ng/ml) produced in our laboratory using an insect expression system [20,21] and then incubated for 24 or 48 h with CMRL supplemented with 0.5% vol./vol. FCS in the presence or absence of a cytokine cocktail (IL-1β, 0.4 ng/ml; TNFα, 4 ng/ml; IFN-γ, 2 ng/ml) (Peprotech, Mexico DF, Mexico) and either vehicle or rhPRL (300 ng/ml).…”

Section: Methodsmentioning

confidence: 99%

Recombinant human prolactin promotes human beta cell survival via inhibition of extrinsic and intrinsic apoptosis pathways

et al. 2011

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Aims/hypothesis Transplantation of pancreatic islets constitutes a promising alternative treatment for type 1 diabetes. However, it is limited by the shortage of organ donors. Previous results from our laboratory have demonstrated beneficial effects of recombinant human prolactin (rhPRL) treatment on beta cell cultures. We therefore investigated the role of rhPRL action in human beta cell survival, focusing on the molecular mechanisms involved in this process. Methods Human pancreatic islets were isolated using an automated method. Islet cultures were pre-treated in the absence or presence of rhPRL and then subjected to serum starvation or cytokine treatment. Beta cells were labelled with Newport green and apoptosis was evaluated using flow cytometry analysis. Levels of BCL2 gene family members were studied by quantitative RT-PCR and western blot. Caspase-8, -9 and -3 activity, as well as nitric oxide production, were evaluated by fluorimetric assays. ResultsThe proportion of apoptotic beta cells was significantly lowered in the presence of rhPRL under both cell death-induced conditions. We also demonstrated that cytoprotection may involve an increase of BCL2/BAX ratio, as well as inhibition of caspase-8, -9 and -3. Conclusions/interpretation Our study provides relevant evidence for a protective effect of lactogens on human beta cell apoptosis. The results also suggest that the improvement of cell survival may involve, at least in part, inhibition of cell death pathways controlled by the BCL2 gene family members. These findings are highly relevant for improvement of the islet isolation procedure and for clinical islet transplantation.

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Section: Methodsmentioning

confidence: 99%

Recombinant human prolactin promotes human beta cell survival via inhibition of extrinsic and intrinsic apoptosis pathways

et al. 2011

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“…The biological activity of hormones produced in Pichia pastoris recombinant was determined by their ability to promote proliferation of the Nb2 cell line, which comes from rat lymphoma (54,55). GHs were also tested for their somatogenic activity in the adipocyte differentiation model based on the glyceraldehyde 3-phosphate dehydrogenase (GPDH) assay.…”

Section: Lactogenic Activity Bioassaymentioning

confidence: 99%

Genetic Engineering and Biotechnology of Growth Hormones

Ascacio-Martínez¹,

Barrera-Saldaña²

2012

Genetic Engineering - Basics, New Applications and Responsibilities

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“…The mitogenic potency of fractionated PRL (bioPRL) was evalu ated on Nbj rat lymphoma cells, as described by Lawson et al [36], with modification [33]. Diluted pituitary extracts and plasma were assayed in 10.0-and 25.0-pl aliquots, respectively.…”

Section: Mb; In Vitro Bioassaymentioning

confidence: 99%

Size and Charge Heterogeneity of Pituitary and Plasma Prolactin in the Male Rat

Briski

Swanson²,

Sylvester³

1996

Neuroendocrinology

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Although pituitary prolactin (PRL) exhibits size and charge variability, questions concerning the structural heterogeneity and biological potency of hormonal forms secreted in vivo remain. In the present studies, monomeric PRL in male rat pituitaries and plasma was fractionated by Sephacryl S-100 size exclusion chromatography and aqueous chromatofocusing to resolve size and charge forms under conditions compatible with optimum preservation of biological activity. Individual hormonal variants were subsequently evaluated for their ability to stimulate the growth of PRL receptor-bearing rat lymphoma cells in vitro. Pituitary elution profiles contained several cross-reactive size variants ranging from 30.4 to 21.5 kD in M_r; major hormonal peaks were eluted at 25.6, 24.3, and 23.6 kD. Multiple size forms of PRL were also detected in plasma profiles, with predominant peaks eluting between 26.5 and 21.5 kD M_Γ in size. Mean B/I ratios, established as an index of relative biopotency, varied significantly between size variants obtained from pituitary and plasma. Pituitary PRL size variants of 27.1 24.3, and 21.5 kD exhibited greatest potency in the in vitro bioassay, whereas the 25.6- and 23.6-kD forms were least potent under these conditions. Of the PRL size variants detected in peripheral plasma, those of 24.3 and 21.5 kD size were characterized by highest mean B/I ratios. Pituitary 24.3-kD PRL was chromatofocused as five charge variants of pI 5.34, 5.31, 5.26, 5.20, and 5.14; only some of these isomers are apparently secreted in vivo, since pI values for plasma charge isomers ranged from 5.26 to 5.14. Charge isomers of pituitary 24.3-kD PRL exerted variable mitogenicity in the Nb₂ in vitro bioassay. The highest mean B/I ratio was associated with the relatively basic 24.3-kD isomer of pI 5.31; the relative biopotency of more acidic charge isomers was progressively diminished with increasing acidic charge. In summary, the present findings show that both size and charge variants of male rat pituitary PRL exhibit differential biopotency in vitro, indicative of their functional heterogeneity. The demonstration of multiple hormonal forms in peripheral plasma suggests that the net biopotency of circulating PRL reflects the sum of activity of structurally and functionally diverse molecules.

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Rat lymphoma cell bioassay for prolactin: Observations on its use and comparison with radioimmunoassay

Cited by 50 publications

References 7 publications

Recombinant human prolactin promotes human beta cell survival via inhibition of extrinsic and intrinsic apoptosis pathways

Recombinant human prolactin promotes human beta cell survival via inhibition of extrinsic and intrinsic apoptosis pathways

Genetic Engineering and Biotechnology of Growth Hormones

Size and Charge Heterogeneity of Pituitary and Plasma Prolactin in the Male Rat

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