New 2,6,9-trisubstituted adenines as adenosine receptor antagonists: a preliminary SAR profile

A new series of 2,6,9-trisubstituted adenines (5–14) have been prepared and evaluated in radioligand binding studies for their affinity at the human A₁, A_2A and A₃ adenosine receptors and in adenylyl cyclase experiments for their potency at the human A_2B subtype. From this preliminary study the conclusion can be drawn that introduction of bulky chains at the N ⁶ position of 9-propyladenine significantly increased binding affinity at the human A₁ and A₃ adenosine receptors, while the presence of a chlorine atom at the 2 position resulted in a not univocal effect, depending on the receptor subtype and/or on the substituent present in the N ⁶ position. However, in all cases, the presence in the 2 position of a chlorine atom favoured the interaction with the A_2A subtype. These results demonstrated that, although the synthesized compounds were found to be quite inactive at the human A_2B subtype, adenine is a useful template for further development of simplified adenosine receptor antagonists with distinct receptor selectivity profiles.     

ICOS Gene Polymorphisms (IVS1 + 173 T/C and c. 1624 C/T) in Primary Sjögren’s Syndrome Patients: Analysis of ICOS Expression

Background: Primary Sjögren’s syndrome (pSS) is a systemic autoimmune disease, which affects exocrine glands. T cell activation is a trigger mechanism in the immune response. Hyperreactivity of T cells and antibody production are features in pSS. ICOS can be critical in the pathogenesis of pSS. Methods: A total of 134 pSS patients and 134 control subjects (CS) were included. Genotyping was performed by PCR-RFLP. ICOS mRNA expression was quantified by real-time PCR, and CD4+ ICOS+ T cells were determined by flow cytometry. Results: The ICOS IVS1 + 173 T>C polymorphisms were not associated with susceptibility to pSS (p = 0.393, CI = 0.503–1.311). However, the c.1624 C>T polymorphism was associated with a reduction in the risk of development of pSS (p = 0.015, CI = 0.294–0.884). An increase in ICOS mRNA expression in patients was observed (3.7-fold). Furthermore, pSS patients showed an increase in membranal-ICOS expression (mICOS). High expression of mICOS (MFI) was associated with lymphocytic infiltration. Conclusions: The IVS1 + 173 polymorphism is not a genetic marker for the development of pSS, while c.1624 T allele was associated with a low risk. However, elevated mICOS expression in pSS patients with high lymphocytic infiltration was found. ICOS may have an important role in the immunopathogenesis of pSS and should be analyzed in T cell subsets in pSS patients as a possible disease marker.     

Isolation and characterization of pLS 1 plasmid mutants with increased copy numbers

Abstract Streptococcus pneumoniae genetic systems designed for isolation of plasmid mutants with copy-up phenotypes have been developed. The target plasmids have the pLS1 replicon, and two different strategies have been followed: (i) selection of clones exhibiting augmented resistance to antibiotics, or (ii) obligatory co-existence of incompatible plasmids. We have isolated 23 plasmid mutants exhibiting increased number of copies. All the mutations corresponded to four different alleles of the copG gene of plasmid pLS1. These strategies could be used with other plasmids.     

Phyllachora species infecting maize and other grass species in the Americas represents a complex of closely related species

The genus Phyllachora contains numerous obligate fungal parasites that produce raised, melanized structures called stromata on their plant hosts referred to as tar spot. Members of this genus are known to infect many grass species but generally do not cause significant damage or defoliation, with the exception of P. maydis which has emerged as an important pathogen of maize throughout the Americas, but the origin of this pathogen remains unknown. To date, species designations for Phyllachora have been based on host associations and morphology, and most species are assumed to be host specific. We assessed the sequence diversity of 186 single stroma isolates collected from 16 hosts representing 15 countries. Samples included both herbarium and contemporary strains that covered a temporal range from 1905 to 2019. These 186 isolates were grouped into five distinct species with strong bootstrap support. We found three closely related, but genetically distinct groups of Phyllachora are capable of infecting maize in the United States, we refer to these as the P. maydis species complex. Based on herbarium specimens, we hypothesize that these three groups in the P. maydis species complex originated from Central America, Mexico, and the Caribbean. Although two of these groups were only found on maize, the third and largest group contained contemporary strains found on maize and other grass hosts, as well as herbarium specimens from maize and other grasses that include 10 species of Phyllachora. The herbarium specimens were previously identified based on morphology and host association. This work represents the first attempt at molecular characterization of Phyllachora species infecting grass hosts and indicates some Phyllachora species can infect a broad range of host species and there may be significant synonymy in the Phyllachora genus.     

Cytochemical Localization of Calcium in Prefusion Myoblasts from the Chick Embryo Myotome

Myoblast fusion is a Ca²⁺-dependent process. The aim of this report was to study the localization of Ca²⁺ in prefusion myoblasts from the brachial somites of chick embryos (51–108h of incubation), using the potassium pyroantimonate cytochemical method. When observed under a transmission electron microscope, electron-dense precipitates of Ca²⁺-antimonate were found in the basement membrane of the myotome, which separates the myotome from the adjacent mesenchyma. Within myoblasts, triads and sarcoplasmic reticulum associated with the first newly formed sarcomeres were observed, but a T-tubule network was not found. Moreover, Ca²⁺-antimonate precipitates were not observed in structures resembling T-tubules or sarcoplasmic reticulum. The results suggest that sarcomerogenesis and sarcoplasmic reticulum development occur simultaneously and that prefusion myoblasts have neither a T-tubule network nor Ca²⁺ deposits on sarcoplasmic reticulum. Small Ca²⁺ pools were found in the myoblast nuclei, cytoplasmic vesicles and mitochondrias. Ca²⁺-antimonate precipitates periodically distributed at the cell periphery, close to the cell membrane, were observed. These precipitates could represent internal Ca²⁺ stores located in the peripheral couplings and it is proposed that these pools of Ca²⁺ could be mobilized before fusion, leading to the increase in free intracellular Ca²⁺ that precedes myoblast fusion.     

Constituents of Quinchamalium majus with potential antitubercular activity

Antitubercular bioassay-guided fractionation of the dichloromethane extracts of the above-ground biomass and roots of Quinchamalium majus led to the identification of six known constituents, betulinic acid (1), daucosterol (2), 5,7-dihydroxyflavone (3), oleanolic acid (4), (-)-2S-pinocembrin (5), and ursolic acid (6), for the first time in this species. Their chemical structures were determined on the basis of spectroscopic evidence and chemical transformation methods. All of these compounds along with additional 11 analogues were evaluated for their antitubercular potential against Mycobacterium tuberculosis in a microplate alamar blue assay, and the primary structure-activity relationships (SARs) for 4 and 6 were discussed. In addition, all the isolates were tested for cytotoxicity against African green monkey Vero cells in order to evaluate for their selectivity potential.