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Targeting ulcerative colitis by suppressing glucose uptake with ritonavir.

ABSTRACT: Glucose is the preferred source of energy in activated inflammatory cells. Glucose uptake into the cell is ensured by a family of glucose uptake transporters (GLUTs), which have been identified as off-target molecules of the HIV protease inhibitor ritonavir. In this study, we examined the effect of ritonavir on inflammation in vitro and in vivo Peripheral blood mononuclear cells (PBMCs) were activated with anti-CD3 in the presence or absence of ritonavir and analyzed by flow cytometric analysis. Frequencies of CD4+ cells were significantly affected by ritonavir (CD69+ P=3E-05; CD134 P=4E-06; CD25+ P=E-07; central memory P=0.02; effector P=6E-03; effector memory P=6E-05). To corroborate that inflammation has a metabolic effect in vivo, a mouse model was used that is based on immunocompromised NOD-scid IL-2R? ?null mice reconstituted with PBMCs from patients with ulcerative colitis (UC). Inflammation had a significant effect on amino acid (AA) levels (Glu P=1E-07, Asp P=1E-04). Principal component analysis (PCA) discriminated between unchallenged and challenged groups. Finally, the efficacy of ritonavir was tested in the same mouse model. Dependent variables were clinical and histological scores, frequencies of human leukocytes isolated from spleen and colon, and levels of AA in sera of mice. Mice benefited from treatment with ritonavir as indicated by significantly decreased colon (P=7E-04) and histological (P=1E-04) scores, frequencies of M2 monocytes (CD14+ CD163; P=0.02), and Glu levels (P=2E-05). PCA discriminated between control and challenged groups (P=0.026). Thus, inhibition of glucose uptake might be a promising therapeutic intervention point for active UC.

SUBMITTER: Jodeleit H

PROVIDER: S-EPMC6262818 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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Targeting ulcerative colitis by suppressing glucose uptake with ritonavir.

Jodeleit Henrika H Al-Amodi Omar O Caesar Janina J Villarroel Aguilera Christina C Holdt Lesca L Gropp Roswitha R Beigel Florian F Siebeck Matthias M

Disease models & mechanisms 20181121 11

Glucose is the preferred source of energy in activated inflammatory cells. Glucose uptake into the cell is ensured by a family of glucose uptake transporters (GLUTs), which have been identified as off-target molecules of the HIV protease inhibitor ritonavir. In this study, we examined the effect of ritonavir on inflammation <i>in vitro</i> and <i>in vivo</i> Peripheral blood mononuclear cells (PBMCs) were activated with anti-CD3 in the presence or absence of ritonavir and analyzed by flow cytome ...[more]

PMID: 30322872

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Project description:BackgroundThe production of integral membrane spanning proteins (IMP's) constitutes a bottleneck in pharmaceutical development. It was long considered that the state-of-the-art was to produce the proteins as inclusion bodies using a powerful induction system. However, the quality of the protein was compromised and the production of a soluble protein that is incorporated into the membrane from which it is extracted is now considered to be a better method. Earlier research has indicated that a slower rate of protein synthesis might overcome the tendency to form inclusion bodies. We here suggest the use of a set of E. coli mutants characterized by a slower rate of growth and protein synthesis as a tool for increasing the amount of soluble protein in high- throughput protein production processes.ResultsA set of five IMP's was chosen which were expressed in three mutants and the corresponding WT cell (control). The mutations led to three different substrate uptake rates, two of which were considerably slower than that of the wild type. Using the mutants, we were able to express three out of the five membrane proteins. Most successful was the mutant growing at 50% of the wild type growth rate. A further effect of a low growth rate is a low acetic acid formation, and we believe that this is a possible reason for the better production. This hypothesis was further supported by expression from the BL21(DE3) strain, using the same plasmid. This strain grows at a high growth rate but nevertheless yields only small amounts of acetic acid. This strain was also able to express three out of the five IMP's, although at lower quantities.ConclusionsThe use of mutants that reduce the specific substrate uptake rate seems to be a versatile tool for overcoming some of the difficulties in the production of integral membrane spanning proteins. A set of strains with mutations in the glucose uptake system and with a lower acetic acid formation were able to produce three out of five membrane proteins that it was not possible to produce with the corresponding wild type.

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Targeting ulcerative colitis by suppressing glucose uptake with ritonavir.

Publications

Targeting ulcerative colitis by suppressing glucose uptake with ritonavir.

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