Figure 4.

Serine/glycine restriction reduced intracellular glutathione levels that are necessary for hMPC proliferation.

A) The glutathione synthesis pathway with genes upregulated after serine/glycine restriction according to RNA-seq is in blue. CBS, cystathionine β-synthase; CTH, cystathionine gamma-lyase; GCLC, glutamate-cysteine ligase catalytic subunit; GSS, glutathione synthetase.

B) Total intracellular glutathione levels in hMPCs after 5 days in serine/glycine restricted or replete media.

C) Ratio of reduced to oxidized intracellular glutathione levels in hMPCs after 5 days in serine/glycine restricted or replete media. GSH, reduced glutathione; GSSG, oxidized glutathione.

D) ROS in hMPCs after 5 days in serine/glycine restricted or replete media.

E) Total intracellular glutathione levels in hMPCs after 5 days of serine/glycine restriction with or without 10 μM of cell permeable glutathione ethyl ester (GSHee).

F) ROS in hMPCs after 5 days of serine/glycine restriction with or without 10 μM of GSHee.

G) Live cell count after 5 days of serine/glycine restriction with or without GSHee (10 μM).

H) Total intracellular glutathione levels in hMPCs after 5 days of serine/glycine restriction or 1000 μM of serine/glycine and 100 μM of glutathione inhibitor l-buthionine sulfoximine (GSHi).

I) Live cell count after 5 days of hMPCs cultured in serine/glycine replete media, serine/glycine restricted media, or a combination of serine/glycine replete media and 100 μM of GSHi. ∗p = 0.05 and ∗∗p < 0.01.

All the experiments were repeated with hMPCs derived from the same 5 donors. P values or marks of significance are indicated in the appropriate graphs.