1. Volume 6, Issue 3, Pages 683-692 (September 2000)
In the Absence of Extrinsic Signals, Nutrient Utilization by Lymphocytes Is Insufficient to Maintain Either Cell Size or Viability 
Jeffrey C. Rathmell, Mathew G.Vander Heiden, Marian H. Harris, Kenneth A. Frauwirth, Craig B. Thompson 
Molecular Cell 
Volume 6, Issue 3, Pages (September 2000)
DOI: /S (00)

2. Figure 1 Bcl-XL Expression Correlates with Survival but Not Cell Size
(A) Western blot of control (Neo) and two Bcl-XL transfected FL5.12 clones (Bcl-XL 1E1 and Bcl-XL 4.1) was probed for expression of Bcl-XL.
(B) Survival curves were measured by propidium iodide (PI) exclusion following removal of IL-3.
(C) Mean forward light scatter (a measure of cell size) of FL5.12 cells was determined flow cytometrically upon growth factor withdrawal. Due to extensive death, the control clone size is shown only to 24 hr. Average and standard deviations of triplicates are shown.
Molecular Cell 2000 6, DOI: ( /S (00) )

3. Figure 3 Extent of Atrophy Determines Cell Cycle Reentry Time
Cells were growth factor withdrawn for various periods, then IL-3 was added back and the rate of BrdU incorporation was measured flow cytometrically. Means and standard deviations of triplicates are shown.
Molecular Cell 2000 6, DOI: ( /S (00) )

4. Figure 2
In Contrast to Bcl-XL, Growth Factors Increase Cellular ATP Levels, Promote Survival in a Glucose-Dependent Manner, and Induce glut1 Expression
(A) ATP and ADP content of FL5.12 clones were measured after growth factor withdrawal and corrected by the percent living cells. Curves are drawn to the point where cell viability became less than 10%.
(B) Northern blots prepared from Neo and Bcl-XL 4.1 clones grown in the presence of IL-3, 14 hr in the absence of IL-3, and 3 hr after readdition of IL-3 to 14 hr starved cells were hybridized with rat glut1 cDNA. The 18 and 28S rRNA are shown.
(C and D) Viabilities of FL5.12 Neo and Bcl-XL 4.1 clones were determined in high (11 mM) and low glucose in the presence (C) or absence (D) of IL-3. Low glucose was 0.11 mM in (C) and mM in (D). Mean percent survival and standard deviations for triplicate samples are shown.
Molecular Cell 2000 6, DOI: ( /S (00) )

5. Figure 4 Neglected Primary T Cells Atrophy over Time
Purified T cells were cultured in complete media without stimulation for 3 days.
(A) Cell viability was measured by PI exclusion.
(B) Mean forward light scatter of viable cells was measured flow cytometrically. Means and standard deviations of triplicates are shown.
Molecular Cell 2000 6, DOI: ( /S (00) )

6. Figure 5
Bcl-XL Transgenic T Cells Are Increased in Number but Reduced in Size
(A) Western blot of T cells from control (Non-tg) and two Bcl-XL transgenic lines (pSV40-Bcl-XL-tg and pLck-Bcl-XL-tg) was probed for Bcl-XL expression.
(B) CD44lo T cells from control and transgenic lines were analyzed flow cytometrically to determine forward light scatter.
(C) Averages and standard deviations for T cell numbers and mean forward light scatter of splenic CD4 or CD8 single positive CD44lo T cells are shown for five nontransgenic, three pSV40-Bcl-XL transgenic, and five pLck-Bcl-XL transgenic mice. Statistical significance was determined by two-tailed Student's t test.
Molecular Cell 2000 6, DOI: ( /S (00) )

7. Figure 6
Bcl-XL Transgenic T Cells Are Less Metabolically Active, Maintain Lower Energy Stores than Control T Cells, and Have Retarded Cell Cycle Entry
(A) Purified T cells from nontransgenic and pLck-Bcl-XL transgenic mice were stained with rhodamine 123 and analyzed flow cytometrically.
(B) ATP and ADP contents of purified T cells were determined from five nontransgenic and five pLck-Bcl-XL transgenic mice.
(C) Purified T cells from nontransgenic and pLck-Bcl-XL transgenic mice were cultured in high (11 mM) or low (0.011 mM) glucose. Cell viabilities were determined by PI exclusion and are expressed as the percent viable in low glucose divided by the percent viable in high glucose.
(D) Purified T cells from nontransgenic and pLck-Bcl-XL transgenic mice were stimulated with anti-CD3 and anti-CD28. BrdU was added to the media at the start of some cultures and given in 1 and 4 hr pulses 72 hr after the start of others. The percentage of cells that had incorporated BrdU was determined flow cytometrically. Means and standard deviations of triplicates are shown. Statistical significance was determined by a two-tailed Student's t test.
Molecular Cell 2000 6, DOI: ( /S (00) )

8. Figure 7
Availability of T Cell Receptor Stimulation Regulates Cell Size and glut1 Expression
(A) T cells were purified from anti-HY TCR/pSV40-Bcl-XL double transgenic mice, analyzed flow cytometrically to determine initial cell size (Pre-Transfer), and adoptively transferred into congenic hosts. The size of adoptively transferred anti-HY T cells 1 week later was determined flow cytometrically by measuring forward light scatter of Thy1.2+ CD44lo idiotype+ T cells. Means and standard deviations are shown from triplicate mice in each group. Statistical significance was determined by two-tailed Student's t test.
(B) Northern blots prepared from non-transgenic T cells that had been stimulated for 0, 2, 6, or 20 hr, and T cells cultured without stimulation for 20 hr were hybridized with rat glut1 cDNA. The 18 and 28S rRNA are shown.
Molecular Cell 2000 6, DOI: ( /S (00) )

9. Figure 8 Atrophy and Loss of Normal Cells Can Occur In Vivo
(A and B) Purified T cells from Thy1.1 expressing nontransgenic (A) or pLck-Bcl-XL transgenic (B) mice were adoptively transferred into nontransgenic or pLck-Bcl-XL transgenic hosts. The mean forward light scatter and number of adoptively transferred T cells 1 week after transfer were determined flow cytometrically. Means and standard deviations are shown from five mice in each recipient group of nontransgenic mice and four mice in each recipient group of pLck-Bcl-XL-transgenic mice. Statistical significance was determined by two-tailed Student's t test.
N.S., insignificant difference.
Molecular Cell 2000 6, DOI: ( /S (00) )