Leigh Syndrome Mouse Model Can Be Rescued by Interventions that Normalize Brain Hyperoxia, but Not HIF Activation.

Leigh syndrome is a devastating mitochondrial disease for which there are no proven therapies. We previously showed that breathing chronic, continuous hypoxia can prevent and even reverse neurological disease in the Ndufs4 knockout (KO) mouse model of complex I (CI) deficiency and Leigh syndrome. Here, we show that genetic activation of the hypoxia-inducible factor transcriptional program via any of four different strategies is insufficient to rescue disease. Rather, we observe an age-dependent decline in whole-body oxygen consumption. These mice exhibit brain tissue hyperoxia, which is normalized by hypoxic breathing. Alternative experimental strategies to reduce oxygen delivery, including breathing carbon monoxide (600 ppm in air) or severe anemia, can reverse neurological disease. Therefore, unused oxygen is the most likely culprit in the pathology of this disease. While pharmacologic activation of the hypoxia response is unlikely to alleviate disease in vivo, interventions that safely normalize brain tissue hyperoxia may hold therapeutic potential.

Elimination of a human T-cell region in staphylokinase by T-cell screening and computer modeling.

Staphylokinase is a potent highly fibrin-selective thrombolytic agent, but it induces a humoral immune response in most treated patients. Staphylokinase-specific T-lymphocytes can be found in normal healthy individuals, from whom a large panel of staphylokinase-specific T-cells were cloned. The staphylokinase amino acid sequence 71-87 was widely recognized, as it induced proliferation of T-cell clones isolated from 90% of the donors. Computer modeling of this area, threaded as 11-mer peptides within the peptide-binding groove of the major HLA-DR alleles, indicated two putative partially overlapping binding sequences. The region-(71-87)-specific T-cell clones recognized either one or the other minimal peptide, confirming that both sequences could be functional T-cell epitopes. Furthermore, to guide the mutagenesis to eliminate T-cell reactivity, the contribution of each residue to the HLA-DR-anchoring and T-cell receptor exposure was evaluated for both binding motifs. Computer calculations combined with functional assays resulted in the design of staphylokinase-variants, including 2 to 4 amino acid substitutions in the region 71-87. These variants were no longer recognized by the region-(71-87)-specific T-cell clones, and importantly no new staphylokinase-variant-specific cellular immune response could be measured.