Growth of human embryonic fibroblasts at clonal density: Concordance with results from mass cultures

Abstract

In the past, it has been difficult to grow human diploid fibroblast cells at clonal densities. Newly devised cell culture media and rigorously controlled environmental conditions have greatly increased the ease with which such cells can be cloned. The present work was undertaken to determine whether, under appropriate conditions, diploid fibroblast cells from human embryonic lung, grow as well at clonal densities as in mass culture. The parameters studied were: (1) population doubling time, (2) in vitro proliferative capacity, (3) attachment, (4) percentage of non-dividing cells. In all cases essentially the same results were obtained for cultures at clonal densities and mass cultures. These results indicate that the behavior of these types of cells in clonal culture can be used to infer the behavior of individual cells and clones within a mass culture.

Catabolic stress induces features of chondrocyte senescence through overexpression of caveolin 1: Possible involvement of caveolin 1–induced down‐regulation of articular chondrocytes in the pathogenesis of osteoarthritis

“…The end of the replicative lifespan was defined by failure of the population to double after 4 weeks in mass cultures. The in vitro lifespan (remaining replicative capacity) was expressed as population doubling (PD) up to cellular senescence (25). The population doublings were calculated with the equation PD ϭ log 10 (N/N 0 ) ϫ 3.33, where N is the number of cells at the end of the experiment and N 0 is the number of cells at the beginning of the experiment (26).…”

Telomerized presenescent osteoblasts prevent bone mass loss in vivo

“…The in vitro lifespan (remaining replicative capacity) was expressed as population doublings up to cellular senescence. 5,26 Telomerase assay Presenescent osteoblasts from donors, transfected with either the hTERT cDNA-expressing plasmid or control plasmid, were treated with ice-cold hypotonic detergent lysis buffer (10 6 cells/100 ml: 0.5% CHAPS (Sigma)), incubated on ice for 30 min, and centrifuged at 12 000 g for 20 min at 41C. The supernatant was stored at À801C.…”

The Concept of Telomeric Non-Reciprocal Recombination (TENOR) Applied to Human Fibroblasts Grown in Serial Cultures: Concordance with Genealogical Data

“…If cultures were continuously kept in a logarithmic growth state, there was initially enhanced proliferation activity. 53,54 This is explainable by population genetic tools. If telomeric length polymorphism at each chromosome end in a fibroblast culture is considered as an analogue to the allele frequencies at a single gene locus in a natural population of individuals, one can use the concept of genetic drift.…”

“…2,3,7,9,26,38,44,45 As shown for a great number of different fibroblast strains, there is a progressive increase in replicatively senescent cells until the culture cannot be propagated further. 2,[4][5][6][7][8][9][10][11][12][13][14][15]46,47 It has been repeatedly demonstrated that the onset of that terminal state is clearly related to the replicative age, that is, to mitotic time, of the culture and not to chronological time, [48][49][50][51] and that cell proliferation capacities are not adversely affected by growth at low cell density [52][53][54] nor by enzymatic treatment at passaging. 55 Therefore, the consumption of an intrinsically determined proliferation capacity by divisions represents the final replicative boundary which human fibroblasts cannot continuously bypass.…”