Lasioglossins LLIII affect the morphogenesis of Candida albicans and reduces the duration of experimental vaginal candidiasis in mice

Lasioglossins are a group of peptides with identified antimicrobial activity. The inhibitory effects of two synthetic lasioglossin derivatives, LLIII and D‐isomeric variant LLIII‐D, on morphological changes in Candida albicans in vitro and the effect of local administration of LLIII during experimental murine candidiasis were investigated. C. albicans blastoconidia were grown in the presence of lasioglossin LLIII or LLIII‐D at concentrations of 11.5 μM and 21 μM, respectively, for 1, 2 and 3 days and their viability determined by flow cytometry using eosin Y staining. Morphological changes were examined by light and fluorescent microscopy. The Candida‐inhibitory effect of daily intravaginal administration of 0.7 or 1.4 μg of LLIII was assessed in mice with experimentally‐induced vaginal candidiasis. LLIII and LLIII‐D lasioglossins exhibited candidacidal activity in vitro (>76% after 24 hr and >84% after 48 hr of incubation). After 72 hr incubation of Candida with low concentration of lasioglossins, an increase in viability was detected, probably due to a Candida antimicrobial peptides evasion strategy. Furthermore, lasioglossins inhibited temperature‐induced morphotype changes toward hyphae and pseudohyphae with sporadic occurrence of atypical cells with two or enlarged nuclei, suggesting interference with mitosis or cytokinesis. Local application of LLIII reduced the duration of experimental candidiasis with no evidence of adverse effects. Lasioglossin LLIII is a promising candidate for development as an antimicrobial drug for treating the vaginal candidiasis.

     

Expression of subunits of the 19S complex and of the PA28 activator in rat skeletal muscle

A precise knowledge of the role of subunits of the 19S complex and the PA28 regulator, which associate with the 20S proteasome and regulate its peptidase activities, may contribute to design new therapeutic approaches for preventing muscle wasting in human diseases. The proteasome is mainly responsible for the muscle wasting of tumor-bearing and unweighted rats. The expression of some ATPase (MSS1, P45) and non ATPase (P112-L, P31) subunits of the 19S complex, and of the two subunits of the PA28 regulator, was studied in such atrophying muscles. The mRNA levels for all studied subunits increased in unweighted rats, and analysis of MSS1 mRNA distribution profile in polyribosomes showed that this subunit entered active translation. By contrast, only the mRNA levels for MSS1 increased in the muscles from cancer rats. Thus, gene expression of the proteasome regulatory subunits depends on a given catabolic state. Torbafylline, a xanthine derivative which inhibits tumor necrosis factor production, prevented the activation of protein breakdown and the increased expression of 20S proteasome subunits in cancer rats, without reducing the elevated MSS1 mRNA levels. Thus, the increased expression of MSS1 is regulated independently of 20S proteasome subunits, and did not result in accelerated proteolysis.     

Mitochondrial Complex I superoxide production is attenuated by uncoupling

Complex I, i.e. proton-pumping NADH:quinone oxidoreductase, is an essential component of the mitochondrial respiratory chain but produces superoxide as a side-reaction. However, conditions for maximum superoxide production or its attenuation are not well understood. Unlike for Complex III, it has not been clear whether a Complex I-derived superoxide generation at forward electron transport is sensitive to membrane potential or protonmotive force. In order to investigate this, we used Amplex Red for H(2)O(2) monitoring, assessing the total mitochondrial superoxide production in isolated rat liver mitochondria respiring at state 4 as well as at state 3, namely with exclusive Complex I substrates or with Complex I substrates plus succinate. We have shown for the first time, that uncoupling diminishes rotenone-induced H(2)O(2) production also in state 3, while similar attenuation was observed in state 4. Moreover, we have found that 5-(N-ethyl-N-isopropyl) amiloride is a real inhibitor of Complex I H(+) pumping (IC(50) of 27 microM) without affecting respiration. It also partially prevented suppression by FCCP of rotenone-induced H(2)O(2) production with Complex I substrates alone (glutamate and malate), but nearly completely with Complexes I and II substrates. Sole 5-(N-ethyl-N-isopropyl) amiloride alone suppressed 20% and 30% of total H(2)O(2) production, respectively, under these conditions. Our data suggest that Complex I mitochondrial superoxide production can be attenuated by uncoupling, which means by acceleration of Complex I H(+) pumping due to the respiratory control. However, when this acceleration is prevented by 5-(N-ethyl-N-isopropyl) amiloride inhibition, no attenuation of superoxide production takes place.     

Synthesis and antitubercular activity of ferrocenyl diaminoalcohols and diamines

A total of 21 ferrocenyl and benzyl diaminoalcohols and diamines were synthesized and evaluated against Mycobacterium tuberculosis H37Rv. Interestingly, ferrocenyl diamines exhibit better activities than ferrocenyl diaminoalcohols.     

PKC zeta controls DNA topoisomerase-dependent human caspase-2 pre-mRNA splicing

Caspase-2 exists as two main isoforms: the caspase-2L long isoform, which is pro-apoptotic, and the caspase-2S short isoform, which may be anti-apoptotic. Topoisomerase inhibitors drive inclusion of exon 9, specific for Casp-2S mRNA, and lower Casp-2L [corrected] mRNA and protein. With cell lines engineered to express various PKC isoforms, we demonstrate that PKC zeta, but not PKCalpha, positively regulates Casp-2S mRNA assembly triggered by topoisomerase inhibitors. In addition, exon 9 inclusion is lowered in mitosis but increased in the G1/S phase. Hence, the control of caspase-2 exon 9 inclusion by topoisomerase inhibitors depends on phosphorylation and/or dephosphorylation events, and on the cell cycle phase.     

Lysosomal and proteasome-dependent proteolysis are differentially regulated by insulin and/or amino acids following feeding in young, mature and old rats

Skeletal muscle proteolysis is inhibited by oral feeding in the young and mature but not in the elderly. However, the proteolytic pathway(s) responsible for the decreased muscle proteolysis in the postprandial (PP) state is (are) unknown in the young. Moreover, muscle proteolysis is inhibited by both insulin (INS) and amino acids (AA) in vitro, but their respective roles on specific proteolytic pathways in vivo remain to be elucidated. The aim of this study was to investigate the respective role of INS and AA on the inhibition of proteolytic pathways in the PP state in skeletal muscles from young, mature and old rats. Rats were fed over 1 h either a 25% (AA+) or a 0% (AA-) amino acid/protein meal. In each nutritional condition, PP insulin secretion was maintained (AA+/INS+ and AA-/INS+) or blocked (AA+/INS- and AA-/INS-) with diazoxide injections. We report that the PP inhibition of proteolysis in young rats was mediated by the increased INS secretion and resulted from a down-regulation of both lysosomal and Ca(2+)-dependent proteolysis. Moreover, our data showed that proteasome activities are inhibited by either INS or AA in mature rats, whereas they become selectively insensitive to AA in old rats. In conclusion, the present work provides direct evidence that the lack of PP regulation of proteasome-dependent proteolysis in old rats resulted from a selective resistance to AA.     

The protein kinases tightly bound to DNA are present in normal tissues and in regenerating liver, but strongly decreased in hepatomas

We have previously described in rat liver two protein kinases tightly bound to DNA, one is serine-specific, the other arginine-specific. In this work we show that both enzymes are present in various rat tissues and in liver from various species. Both kinase specific activities are strongly decreased in methyl-DBA-induced hepatomas and in HTC cells but not in regenerating liver after hepatectomy. This decrease is then not related to cell proliferation.     

Synthesis and antimycobacterial activity of ferrocenyl ethambutol analogues and ferrocenyl diamines

A new series of ferrocenyl diamino alcohols and diamines were synthesized and their inhibitory potencies were probed with Mycobacterium tuberculosis. Interestingly, ferrocenyl diamines 6a and b display significant activities against M. tuberculosis H37Rv.     

Persistence of a vegetation mosaic in a peripheral region: could turbulent medieval history disrupt Holocene continuity of extremely species-rich grasslands?

Fluctuations in intensity of human impact and corresponding vegetation changes have been reported from different parts of Europe for the period from the beginning of the 1st millennium ad to the high Middle Ages. In the Bílé Karpaty mountains (White Carpathians), a region well-known for its biologically valuable ancient grasslands, an extensive spread of woodland could have occurred in the Migration period (4th–6th century) and especially in the Confinium period (11th–12th century), when settling of this border region was legally prohibited. However, Holocene continuity of non-woodland vegetation was suggested as an explanation for the unique species richness of the local grasslands. If this explanation is true, then the turbulent times in medieval history could not have led to complete re-establishment of woodland. To test this idea palaeoecologically, we analysed four new profiles from wetland deposits for pollen, macrofossils and abiotic proxies, and re-dated some old profiles from the area. The results show the continual presence of human impact indicators since the Migration period in the southwest of the Bílé Karpaty, where these unique grasslands occur. Agricultural activities were indicated by pollen of crops, ruderals, weeds and grassland taxa and by macrofossils of fen-grassland plants. Grazing and burning seem to have been the main disturbances during the older period, while mowing of meadows by scythe became more important since the 17th century. Fossil records differed among the sites as a consequence of differences in altitude and disturbance regimes, but converged gradually with time. Despite intensification of human activities, the landscape remained mosaic-like. Indicators of undisturbed woodlands have been detected only in the northeast. Continuous yet perhaps never too intensive disturbances might therefore have maintained the ancient grassland species pool in the long term.