A yeast-based screening assay identifies repurposed drugs that suppress mitochondrial fusion and mtDNA maintenance defects.

Mitochondria continually move, fuse and divide, and these dynamics are essential for the proper function of the organelles. Indeed, the dynamic balance of fusion and fission of mitochondria determines their morphology and allows their immediate adaptation to energetic needs as well as preserving their integrity. As a consequence, mitochondrial fusion and fission dynamics and the proteins that control these processes, which are conserved from yeast to human, are essential, and their disturbances are associated with severe human disorders, including neurodegenerative diseases. For example, mutations in OPA1, which encodes a conserved factor essential for mitochondrial fusion, lead to optic atrophy 1, a neurodegeneration that affects the optic nerve, eventually leading to blindness. Here, by screening a collection of ∼1600 repurposed drugs on a fission yeast model, we identified five compounds able to efficiently prevent the lethality associated with the loss of Msp1p, the fission yeast ortholog of OPA1. One compound, hexestrol, was able to rescue both the mitochondrial fragmentation and mitochondrial DNA (mtDNA) depletion induced by the loss of Msp1p, whereas the second, clomifene, only suppressed the mtDNA defect. Yeast has already been successfully used to identify candidate drugs to treat inherited mitochondrial diseases; this work may therefore provide useful leads for the treatment of optic atrophies such as optic atrophy 1 or Leber hereditary optic neuropathy.

[Effects of Yi Fu Ning soft gelatin capsules on reproductive endocrine-immune function of ovariectomized rats].

OBJECTIVE: To investigate the effect of Yi Fu Ning Soft Gelatin Capsules (YFN) on reproductive endocrine-immune function of ovariectomized rats. METHODS: 60 3-month old female Sprague-Dawley rats were used, 50 of them were ovariectomized and randomly divided into 5 groups: ovariectomizy (OVX) group, OVX with diethylstilbestrol tablets (DT) group, OVX with YFN (high dose, middle dose and low dose) group. The others were sham-operated group. The rats were administrated initially in the 4th week after the operation. After drugs had been given for 12 weeks the rats were sacrificed, blood serum hormone, IL-2 content and T lymphocyte subpopulation were detected with methods radioimmunoassay and flow cytometry (FCM). RESULTS: (1) Compared with sham group, the level of serum E2, Te and P significantly decreased (P < 0.01), FSH, LH content significantly increased; Blood T lymphocyte subpopulation CD3+ cells, CD4+ cells and CD4+/CD8+ ratio significantly decreased, serum IL-2 content also significantly decreased (P < 0.01). (2) Compared with model group, after treated by YFN, the level of serum E2 and P significantly increased (P < 0.01), serum FSH and LH content significantly decreased; T lymphocyte subpopulation CD3+ cells, CD4+ cells and CD4+/CD8+ ratio were improved significantly and serum interleukin-2 (IL-2) content increased significantly. CONCLUSION: YFN can increase serum sexual hormone content,reduce the level of FSH and LH, and improve imbalanced T lymphocyte subpopulation, stimulate IL-2 excretion, which means YFN can regulate inordinate reproductive endocrine-immune network in ovariectomized rats.

(2R*,3S*)-1-[125I]Iodo-2,3-bis(4-hydroxyphenyl)pentane ([125I]iodonorhexestrol) and (2R*,3S*)-1-[77Br]Bromo-2,3-bis(4-hydroxyphenyl)pentane ([77Br]bromonorhexestrol), two gamma-emitting estrogens that show receptor-mediated uptake by target tissues in vivo.

Two gamma-emitting estrogen analogues, (2R*,3S*)-1-[125I]iodo-2,3-bis(4-hydroxyphenyl)pentane ([125I]iodonorhexestrol) and (2R*,3S*)-1-[77Br]bromo-2,3-bis(4-hydroxyphenyl]pentane ([77Br]bromonorhexestrol), have been prepared by halide ion displacement on a labile trifluoromethanesulfonate derivative of a suitably protected precursor, followed by mild acid deprotection. Although halide displacement on a more stable tristrifluoromethanesulfonate derivative was successful, the basic conditions required for deprotection of this precursor resulted in destruction of the products by a base-induced spiroelimination reaction. In immature female rats, both of these halonorhexestrols demonstrated preferential uptake by the uterus that could be blocked selectively by coadministration of a large dose of unlabeled estradiol. In a double label comparison with 16 alpha-[125I]iodo-17 beta-estradiol the uterine uptake of [77Br]bromonorhexestrol was notably less selective. Stability studies in vitro and in vitro have indicated that both iodo- and bromonorhexestrol are quite labile, and this lability compromises the selectivity of their uptake by estrogen target tissues in vivo. p-Hydroxyphenethyl halides are known to be unusually prone to a base-catalyzed solvolysis, via cyclization of the phenolate to a spirocyclohexadienone intermediate. This unusual solvolytic mechanism may contribute to the lability of these halonorhexestrols in vivo.

Estrogen withdrawal in chick oviduct. Characterization of RNA synthesized in isolated nuclei using a mercurated precursor.

1. As a prerequisite for analyzing the effect of estrogen on transcription in chick oviduct, we describe suitable methods for the synthesis (under conditions restricting reinitiation), and isolation of RNA transcripts from oviduct nuclei in vitro, utilizing mercurated UTP (Hg-UTP) as an RNA precursor and chromatography on sulphydryl-Sepharose (SH-Sepharose) to recover mercurated RNA (Hg-RNA). The techniques described include treatment of Hg-RNA with p-hydroxymercuribenzoate, to improve the efficiency of binding to SH-Sepharose, and elution of Hg-RNA from SH-Sepharose after treatment with 60% formamide at 90 degrees C, to eliminate contamination by aggregated nucleic acid. 2. RNA synthesized by endogenous form B RNA polymerase (using either UTP or Hg-UTP as precursor) was recovered in nuclear lysates in the form of 30--85-S heterogeneous RNA . protein complexes, and after removal of protein, was 10--12 S in size. 3. The nature of RNA transcripts synthesized in vitro was examined by hybridization. More than 90% of the RNA was complementary to "unique" DNA sequences, and 50--60% of the hybridized RNA could be competed with homologous, steady-state nuclear RNA, indicating a significant degree with homologous, steady-state nuclear RNA, indicating a significant degree of homology between in vitro transcripts and in vivo RNA. The level of homology was similar whether RNA synthesis was performed in low salt, or in high salt in the presence of heparin. Possible reasons for only partial competition in these experiments are discussed. 4. Withdrawal of estrogen from chicks leads to a 50% reduction in endogenous RNA polymerase activities in nuclei within 48 h. Similar levels of competition with Hg-RNA transcripts for "unique" DNA were obtained using oviduct nuclear RNAs isolated before or after estrogen withdrawal, and even with liver nuclear RNA. Thus, in oviduct, those sequences present in primary transcripts, and analyzed under our experimental conditions, are present in different hormonal states and also in other chick tissues.