Population doubling

A method of assessing the toxicity of implants has been proposed based on the effects on cell ultrastructure in organ cultures, on cell surface characteristics, and cell population doubling times. The effects have been correlated with hemorrhage, fibrosis, and necrosis, respectively (103). Poly-e-caprolactone was stated to give minimal tissue reaction and could not be scored in these tests. 

The recent gains in world population and the pressure they place on natural resources is having an impact on the economics of weed control currently and will have far reaching impact on it in the future. Since the early 1940 s the world s population doubled -from less than 2.3 billion to over 4.6 billion in 1983. The annual rate of growth is about 1.7 percent and recent projections indicate the global population will increase to slightly over 6 billion by 2000. 

Initially, the cells require a period of time, the lag phase, to adjust to the altered environment. No cell multiplication occurs until the individual cells have almost doubled in size. After the adaptation is complete, the cells begin to reproduce exponentially. With each period of cell division, the population doubles. Some cells... 

Figure 26.1 Immortalization of human cells Cells enter replicative senescence at mortality stage 1 (Ml Hayflick limit) after about 60 population doublings (PD). The protein p 16 accumulates in senescent cells. The simian virus 40 (SV40) large T antigen as well as the human papilloma virus (HPV) type 16-E6 and E7 proteins sequester the retinoblastoma protein (Rb) and/or p53 constitutively releases the transcription factor E2F. E2F induces expression proteins required for progression through Gl/S transition, thus the cells escape cell cycle arrest. At mortality stage 2 (M2), transformed cells must overcome senescence and crisis before they are immortalized. This is likely to involve the activation of telomerase either by the introduction of hTERT cDNA or by a genetic change that activates telomerase.

Based on the results from those cell calibration assays, the routine incubation period was set at 3 days. During that interval, some of the cells will proliferate, some will be quiescent but metabolically active, and some will die, which is the case for most primary cell cultures. Because these cells are cultured for periods far shorter than their typical population doubling time (about 6 to 7 days), these cells should not fully adapt to the tissue culture conditions and, therefore, maintain their primary cell phenotype. 

This cultivation method is defined by a limited number of passages or population doublings which must not be exceeded during production. The maximum number of cell doublings, or passage levels, during which the manufacturing process routinely meets the criteria described below, must be stated. 

By this cultivation method the number of passages or population doublings is not restricted from the beginning of production. Criteria for the harvesting as well as for the termination of production have to be defined by the manufacturer. Monitoring is necessary throughout the life of the culture the required frequency and type of monitoring will depend on the nature of the production system and the product. 

In a recent study we analyzed the capacity of HPL to replace FBS in large-scale (chnical) MSC expansions and were able to demonstrate a superior propagation of MSC cultured with HPL (MSCHPL) as compared to MSC derived from FBS-driven cultures (MSCFBS) [23]. Figure 4 illustrates superior MSC proliferation at low plating density and higher population doublings (PDs) with HPL after a culture period of less than 14 days. 

There are also risks associated with the long-term expansion of stem cells in culture, such as transformation and aneuploidy. Such modifications of the intrinsic properties of the cells can potentially lead to the development of malignancy in the patient following transplantation. In the protocol developed by Osiris, MSCs are expanded until the number of adult stem cells has increased over 3000-fold [327297], [327352]. Recent studies have reported that MSCs expanded in culture have an intrinsic propensity towards spontaneous transformation after a certain number of population doublings ( 250) [660856], [660857],... 

If world population doubles in 40 years and world food production also doubles in 40 years,how many people then will be starving each year compared to now ... 

The use of an immobilized biofilm system is recommended, since the oxidation products often are only slowly biodegradable. Biomass, especially slow growing microorganisms, can efficiently be kept in the system using support material. This is especially important in continuously operated systems where the liquid phase is completely mixed. In such a system suspended biomass would be washed out when the hydraulic retention time is less than or equal to the population doubling time (i. e. the reciprocal of the specific growth rate Grady, 1985). In several studies materials such as polyurethane foams (Moerman et al., 1994 Jochimsen, 1997) or quartz sand particles (Stem etal., 1995 and 1996 Heinzle et al, 1995 Saupe and Wiesmann, 1998) have been used for immobilization. 

P2. Pagnan, A., Havel, R. J., Kane, J. P., and Kotite, L., Characterization of very low density lipoproteins containing two electrophoretic populations. Double pre-beta lipoproteinemia and primary dysbetalipoproteinemia. J. Lipid Res. 18, 613-622 (1977). 

Total number of population doublings (PDs). The more population doublings a culture has experienced, the greater number of senescent, or possibly near-senescent cells there will be in that culture. The problem will be that a culture at a low PD number may superficially look the same as a population with a higher PD number, and be growing at the same rate, but might behave quite differently in a biocompatibility assay. 

J. C. Houck and V. K. Sharma, Functional failures of cultured human diploid fibroblasts after continued population doublings, Proc. Soc. Experim. Biol. Med. 137(1), 331 (1971)... 

Metabolically incorporate isotopic label into either control or treatment cell cultures for 5 or 6 population doublings... 

Cells were exposed to drugs for 3-5 days, allowing for at least two population doublings. Cell numbers were estimated by quantification of protein content of fixed cells by methylene blue staining. Data from Ref. 38. 

Cells are randomly plated at day 0 in multiple wells or dishes, so to consent at least triplicate determinations for each time point. The initial plating density will determine the successive time points if cells have been plated at medium to high density (about 10 -10 /cm ), cell numbers can be evaluated starting 18-24 h after plating (at which time a cell number close to the actual number of plated cells is expected), and each day until confluence is attained. Population doublings (PD) can then be calculated using the formula (Kath et al., 1990) ... 

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