Cytogenetic characteristics of a population of Chironomus riparius Meigen 1804 (Diptera, Chironomidae) from a polluted Po river station

A population of Chironomus riparius from a Po river station near Moncalieri (a trace-metal polluted station) was studied. In this population was established a great variability of band structure of polytene chromosomes as well as paracentric heterozygous inversions, deletions, deficiencies, partial breaks, diploid chromosome fragments, and changes in functional activity and appearance of heterochromatin. In arms A through F, some bands had an increased size compared to the standard chromosomic map. Some bands appeared in a heterozygous or normal homozygous state or were amplified. In all arms, many condensed stable bands appeared in the decondensed state when compared to the standard map. Asynaptic zones in arms E and G as well as heterozygous Balbiani rings and NORs were established. Very often the 4th chromosome was almost completely heteropycnotic and looded like a pompon chromosome. For the first time in this species, a high frequency of ectopic pairings of different arms was observed. Telomeric regions involved in ectopic pairings had a granular appearance, as did some centromeres. The hypothesis is advanced that such a high frequency of structural rearrangements could be correlated with genomic distribution of specific mobile elements.     

Nodal-dependent Cripto signaling promotes cardiomyogenesis and redirects the neural fate of embryonic stem cells

The molecular mechanisms controlling inductive events leading to the specification and terminal differentiation of cardiomyocytes are still largely unknown. We have investigated the role of Cripto, an EGF-CFC factor, in the earliest stages of cardiomyogenesis. We find that both the timing of initiation and the duration of Cripto signaling are crucial for priming differentiation of embryonic stem (ES) cells into cardiomyocytes, indicating that Cripto acts early to determine the cardiac fate. Furthermore, we show that failure to activate Cripto signaling in this early window of time results in a direct conversion of ES cells into a neural fate. Moreover, the induction of Cripto activates the Smad2 pathway, and overexpression of activated forms of type I receptor ActRIB compensates for the lack of Cripto signaling in promoting cardiomyogenesis. Finally, we show that Nodal antagonists inhibit Cripto-regulated cardiomyocyte induction and differentiation in ES cells. All together our findings provide evidence for a novel role of the Nodal/Cripto/Alk4 pathway in this process.     

Fetal DNA detection in maternal plasma throughout gestation

The presence of fetal DNA in maternal plasma may represent a source of genetic material which can be obtained noninvasively. We wanted to assess whether fetal DNA is detectable in all pregnant women, to define the range and distribution of fetal DNA concentration at different gestational ages, to identify the optimal period to obtain a maternal blood sample yielding an adequate amount of fetal DNA for prenatal diagnosis, and to evaluate accuracy and predictive values of this approach. This information is crucial to develop safe and reliable non-invasive genetic testing in early pregnancy and monitoring of pregnancy complications in late gestation. Fetal DNA quantification in maternal plasma was carried out by real-time PCR on the SRY gene in male-bearing pregnancies to distinguish between maternal and fetal DNA. A cohort of 1,837 pregnant women was investigated. Fetal DNA could be detected from the sixth week and could be retrieved at any gestational week. No false-positive results were obtained in 163 women with previous embryo loss or previous male babies. Fetal DNA analysis performed blindly on a subset of 464 women displayed 99.4, 97.8 and 100% accuracy in fetal gender determination during the first, second, and third trimester of pregnancy, respectively. No SRY amplification was obtained in seven out of the 246 (2.8%) male-bearing pregnancies. Fetal DNA from maternal plasma seems to be an adequate and reliable source of genetic material for a noninvasive prenatal diagnostic approach.     

Molecular modeling study on potent and selective adenosine A(3) receptor agonists

Adenosine A(3) receptor (A(3)AR) is involved in a variety of key physio-pathological processes and its agonists are potential therapeutic agents for the treatment of rheumatoid arthritis, dry eye disorders, asthma, as anti-inflammatory agents, and in cancer therapy. Recently reported MECA (5'-N-methylcarboxamidoadenosine) derivatives bearing a methyl group in N(6)-position and an arylethynyl substituent in 2-position demonstrated to possess sub-nanomolar affinity and remarkable selectivity for the human A(3)AR, behaving as full agonists of this receptor. In this study, we made an attempt to get a rationalization of the high affinities and selectivities of these molecules for the human A(3)AR, by using adenosine receptor (AR) structural models based on the A(2A)AR crystal structure and molecular docking analysis. Post-docking analysis allowed to evaluate the ability of modeling tools in predicting AA(3)R affinity and in providing interpretation of compound substituents effect on the A(3)AR affinity and selectivity.     

Strategies to obtain stable transgenic plants from non-embryogenic lines: complementation of the nn 1 mutation of the NORK gene in Medicago sativa MN1008

Strategies to introduce genes into non-embryogenic plants for complementation of a mutation are described and tested on tetraploid alfalfa (Medicago sativa). Genes conditioning embryogenic potential, a mutant phenotype, and a gene to complement the mutation can be combined using several different crossing and selection steps. In the successful strategy used here, the M. sativa genotype MnNC-1008(NN) carrying the recessive non-nodulating mutant allele nn ₁ was crossed with the highly embryogenic alfalfa line Regen S and embryogenic hybrid individuals were identified from the F1 progeny. After transformation of these hybrids with the wild-type gene (NORK), an F2 generation segregating for the mutation and transgene were produced. Plants homozygous for the mutant allele and carrying the wild-type NORK transgene could form root nodules after inoculation with Sinorhizobium meliloti demonstrating successful complementation of the nn ₁ mutation.     

The Engineering of a Novel Ligand in gH Confers to HSV an Expanded Tropism Independent of gD Activation by Its Receptors

Herpes simplex virus (HSV) enters cells by means of four essential glycoproteins - gD, gH/gL, gB, activated in a cascade fashion by gD binding to one of its receptors, nectin1 and HVEM. We report that the engineering in gH of a heterologous ligand – a single-chain antibody (scFv) to the cancer-specific HER2 receptor – expands the HSV tropism to cells which express HER2 as the sole receptor. The significance of this finding is twofold. It impacts on our understanding of HSV entry mechanism and the design of retargeted oncolytic-HSVs. Specifically, entry of the recombinant viruses carrying the scFv-HER2–gH chimera into HER2⁺ cells occurred in the absence of gD receptors, or upon deletion of key residues in gD that constitute the nectin1/HVEM binding sites. In essence, the scFv in gH substituted for gD-mediated activation and rendered a functional gD non-essential for entry via HER2. The activation of the gH moiety in the chimera was carried out by the scFv in cis, not in trans as it occurs with wt-gD. With respect to the design of oncolytic-HSVs, previous retargeting strategies were based exclusively on insertion in gD of ligands to cancer-specific receptors. The current findings show that (i) gH accepts a heterologous ligand. The viruses retargeted via gH (ii) do not require the gD-dependent activation, and (iii) replicate and kill cells at high efficiency. Thus, gH represents an additional tool for the design of fully-virulent oncolytic-HSVs retargeted to cancer receptors and detargeted from gD receptors.     

Dimorphic expression of uncoupling protein-3 in golden hamster harderian gland: effects of castration and testosterone administration

Hamster (Mesocricetus auratus) harderian gland (HG) is a dimorphic orbital gland producing a copious lipid secretion. Two cell-types are present in hamster HG, type I in both sexes, type II only in males. In hamster HGs, we found a marked sexual dichotomy in the expression of uncoupling protein-3 (UCP3), a mitochondrial protein carrier, that probably exports fatty acid anions and fatty acid peroxides from the mitochondrial matrix. Following castration and/or testosterone treatment: (1) UCP3 levels correlated with the type II-cell percentage, not with testosterone levels, (2) in male HGs, UCP3 was comparable to female levels at 30 days post-castration (when the type II-cell percentage had fallen from 50 to 5%), although testosterone was already near zero at 15 days (when neither the type II-cell percentage nor the UCP3 level had fallen), and testosterone-replacement therapy prevented these changes. Testosterone-treated females possessed type II cells and a UCP3 level about twofold higher than in control females. Males displayed more intense UCP3 immunohistochemical positivity in type I HG cells than females. Hence, testosterone may indirectly control UCP3 expression by regulating the gland's morphological and lipid dimorphism. Straight-chain fatty acids [found in alkyl diacylglycerols (ADGs) in males] are oxidized predominantly in mitochondria, branched-chain fatty acids (abundant in ADGs in females) predominantly in peroxisomes, so we speculate that the higher UCP3 expression in males reflects greater fatty acid flux in HG mitochondria. This is supported by our finding that in female (not male) HGs, the peroxisome-rich fraction contained alpha-methylacyl-CoA racemase (AMACR), an enzyme important in the beta-oxidation of branched-chain fatty acids.     

IL-22 Restrains Tapeworm-Mediated Protection against Experimental Colitis via Regulation of IL-25 Expression

Interleukin (IL)-22, an immune cell-derived cytokine whose receptor expression is restricted to non-immune cells (e.g. epithelial cells), can be anti-inflammatory and pro-inflammatory. Mice infected with the tapeworm Hymenolepis diminuta are protected from dinitrobenzene sulphonic acid (DNBS)-induced colitis. Here we assessed expulsion of H. diminuta, the concomitant immune response and the outcome of DNBS-induced colitis in wild-type (WT) and IL-22 deficient mice (IL-22^-/-) ± infection. Interleukin-22^-/- mice had a mildly impaired ability to expel the worm and this correlated with reduced or delayed induction of TH2 immunity as measured by splenic and mesenteric lymph node production of IL-4, IL-5 and IL-13 and intestinal Muc-2 mRNA and goblet cell hyperplasia; in contrast, IL-25 increased in the small intestine of IL-22^-/- mice 8 and 12 days post-infection compared to WT mice. In vitro experiments revealed that H. diminuta directly evoked epithelial production of IL-25 that was inhibited by recombinant IL-22. Also, IL-10 and markers of regulatory T cells were increased in IL-22^-/- mice that displayed less DNBS (3 mg, ir. 72h)-induced colitis. Wild-type mice infected with H. diminuta were protected from colitis, as were infected IL-22^-/- mice and the latter to a degree that they were almost indistinguishable from control, non-DNBS treated mice. Finally, treatment with anti-IL-25 antibodies exaggerated DNBS-induced colitis in IL-22^-/- mice and blocked the anti-colitic effect of infection with H. diminuta. Thus, IL-22 is identified as an endogenous brake on helminth-elicited TH2 immunity, reducing the efficacy of expulsion of H. diminuta and limiting the effectiveness of the anti-colitic events mobilized following infection with H. diminuta in a non-permissive host.     

Hypertriglyceridemia Is Independently Associated with Renal,but Not Retinal Complications in Subjects with Type 2 Diabetes: A Cross-Sectional Analysis of the Renal Insufficiency And Cardiovascular Events (RIACE) Italian Multicenter Study

Objective

Atherogenic dyslipidemia seems to play a major role in microvascular complications and in residual microvascular risk after statin therapy, which reduces triglycerides up to 40%. We assessed whether raised TG levels are associated with an increased burden from microvascular complications in patients with type 2 diabetes.

Methods

Subjects from the Renal Insufficiency And Cardiovascular Events (RIACE) Italian Multicentre Study (n=15,773) were divided in 4 groups depending on whether they had plasma triglycerides below (NTG, 67.8%) or above (HTG, 32.2%) 1.7 mmol/L and were (42.4%) or not on (57.6%) statin therapy. Estimated GFR (eGFR) was calculated from serum creatinine, albuminuria was measured by immunonephelometry or immunoturbidimetry, and retinopathy was evaluated by fundus examination.

Results

HTG subjects, either with or without statin, had higher prevalence of albuminuria, reduced eGFR and chronic kidney disease (CKD), especially the albuminuric forms, but not of retinopathy, than NTG subjects. In contrast, cardiovascular disease and advanced DR were more prevalent in subjects on statin than in those not, independently of triglyceride levels. Logistic regression analysis confirmed that HTG, without or with statin, was independently associated with micro and macroalbuminuria, mildly to severely reduced eGFR, and all CKD phenotypes, but not with retinopathy. The adjusted odd ratios for CKD increased linearly for every 0.26 mmol/L increase (approximately one decile) in triglyceride levels. The increase was higher with increasing severity of albuminuria, eGFR loss and CKD phenotype as well as in subjects receiving than in those not receiving statin treatment.

Conclusions

Triglycerides are associated with CKD, but not retinopathy in subjects with type 2 diabetes, independently of statin treatment. These data point to a possible role of hypertriglyceridemia in the development of CKD, though it remains to be demonstrated that diabetic individuals might benefit from triglyceride reduction with statins and eventually with combination therapy with fibrates.

Trial Registration

www.ClinicalTrials.gov NCT00715481