A polymorphism in the growth hormone-releasing hormone receptor gene: clinical significance?

Two forms of the growth hormone-releasing hormone (GHRH) receptor (GHRH-R) exist in terms of a polymorphism at codon 57. The most common allele possesses GCG, coding for Ala. This codon can also be ACG, replacing the Ala with Thr. The present study demonstrates that the latter occurs in about 20% of pituitary somatotrophinomas, removed from patients with acromegaly. Somatotrophinomas possessing the alternative allele respond, on average, more strongly to GHRH in terms of GH secretion in vitro than tumors which are homozygous for the more common allele. The distribution of the two allelic forms of the GHRH-R did not significantly differ between acromegalic and non-acromegalic subjects. Thus, while the alternative allelic forms may, at least partially, contribute to the variable response of serum GH levels to i.v. GHRH observed in acromegalic and normal subjects, it is unlikely that subjects possessing the rarer form containing Thr in place of Ala at residue 57 are at increased risk of developing acromegaly.     

Selective BRAFV600E inhibitor PLX4720, requires TRAIL assistance to overcome oncogenic PIK3CA resistance

Documented sensitivity of melanoma cells to PLX4720, a selective BRAFV600E inhibitor, is based on the presence of mutant BRAF(V600E) alone, while wt-BRAF or mutated KRAS result in cell proliferation. In colon cancer appearance of oncogenic alterations is complex , since BRAF, like KRAS mutations, tend to co-exist with those in PIK3CA and mutated PI3K has been shown to interfere with the successful application of MEK inhibitors. When PLX4720 was used to treat colon tumours, results were not encouraging and herein we attempt to understand the cause of this recorded resistance and discover rational therapeutic combinations to resensitize oncogene driven tumours to apoptosis. Treatment of two genetically different BRAF(V600E) mutant colon cancer cell lines with PLX4720 conferred complete resistance to cell death. Even though p-MAPK/ ERK kinase (MEK) suppression was achieved, TRAIL, an apoptosis inducing agent, was used synergistically in order to achieve cell death by apoptosis in RKO(BRAFV600E/PIK3CAH1047) cells. In contrast, for the same level of apoptosis in HT29(BRAFV600E/PIK3CAP449T) cells, TRAIL was combined with 17-AAG, an Hsp90 inhibitor. For cells where PLX4720 was completely ineffective, 17-AAG was alternatively used to target mutant BRAF(V600E). TRAIL dependence on the constitutive activation of BRAF(V600E) is emphasised through the overexpression of BRAF(V600E) in the permissive genetic background of colon adenocarcinoma Caco-2 cells. Pharmacological suppression of the PI3K pathway further enhances the synergistic effect between TRAIL and PLX4720 in RKO cells, indicating the presence of PIK3CA(MT) as the inhibitory factor. Another rational combination includes 17-AAG synergism with TRAIL in a BRAF(V600E) mutant dependent manner to commit cells to apoptosis, through DR5 and the amplification of the apoptotic pathway. We have successfully utilised combinations of two chemically unrelated BRAF(V600E) inhibitors in combination with TRAIL in a BRAF(V600E) mutated background and provided insight for new anti-cancer strategies where the activated PI3KCA mutation oncogene should be suppressed.     

Some, but not all, retromer components promote morphogenesis of C. elegans sensory compartments

The endings of sensory receptor cells often lie within specialized compartments formed by glial cells. The main sensory organ of Caenorhabditis elegans, the amphid, provides a powerful setting for studying glial compartment morphogenesis. Our previous studies showed that amphid compartment size is controlled by opposing activities of the Nemo-like kinase LIT-1, which promotes compartment expansion, and the Patched-related protein DAF-6, which restricts compartment growth. From a genetic screen for mutations able to suppress the bloated sensory compartments of daf-6 mutants, we identified an allele of the sorting nexin gene snx-1. SNX-1 protein is a component of the retromer, a protein complex that facilitates recycling of transmembrane proteins from the endosome to the Golgi network. We find that snx-1 functions cell autonomously within glia to promote sensory compartment growth, and that SNX-1 protein is enriched near the surface of the sensory compartment. snx-1 interacts genetically with lit-1 and another regulator of compartment size, the Dispatched-related gene che-14. Mutations in snx-3 and vps-29, also retromer genes, can suppress daf-6 defects. Surprisingly, however, remaining retromer components seem not to be involved. Our results suggest that a novel assembly of retromer components is important for determining sensory compartment dimensions.     

Microbial Diversity of Bovine Mastitic Milk as Described by Pyrosequencing of Metagenomic 16s rDNA

Dairy cow mastitis is an important disease in the dairy industry. Different microbial species have been identified as causative agents in mastitis, and are traditionally diagnosed by bacterial culture. The objective of this study was to use metagenomic pyrosequencing of bacterial 16S rRNA genes to investigate bacterial DNA diversity in milk samples of mastitic and healthy dairy cows and compare the results with those obtained by classical bacterial culture. One hundred and thirty-six milk samples were collected from cows showing signs of mastitis and used for microbiological culture. Additionally, 20 milk samples were collected from healthy quarters. Bacterial DNA was isolated from the same milk samples and the 16S rRNA genes were individually amplified and pyrosequenced. Discriminant analysis showed that the groups of samples that were most clearly different from the rest and thus easily discriminated were the normal milk samples from healthy cows and those characterised by culture as Trueperella pyogenes and Streptococcus spp. The mastitis pathogens identified by culture were generally among the most frequent organisms detected by pyrosequencing, and in some cases (Escherichia coli, Klebsiella spp. and Streptococcus uberis mastitis) the single most prevalent microorganism. Trueperella pyogenes sequences were the second most prevalent sequences in mastitis cases diagnosed as Trueperella pyogenes by culture, Streptococcus dysgalactiae sequences were the second most prevalent sequences in mastitis cases diagnosed as Streptococcus dysgalactiae by culture, and Staphyloccocus aureus sequences were the third most prevalent in mastitis cases diagnosed as Staphylococcus aureus by culture. In samples that were aerobic culture negative, pyrosequencing identified DNA of bacteria that are known to cause mastitis, DNA of bacteria that are known pathogens but have so far not been associated with mastitis, and DNA of bacteria that are currently not known to be pathogens. A possible role of anaerobic pathogens in bovine mastitis is also suggested.     

Imprint cytology of vacuum-assisted breast biopsy specimens: a rapid diagnostic tool in non-palpable solid lesions

Objective:  Imprint cytology provides a rapid preliminary diagnosis shortly after the completion of breast biopsy. This study aims to assess the validity of imprint cytology for the pre-operative diagnosis of non-palpable mammographic solid lesions excised by vacuum-assisted breast biopsy (VABB).
Methods:  Seventy-two women with non-palpable Breast Imaging Reporting and Data System 3 and 4 mammographic solid lesions without microcalcifications underwent VABB on the stereotactic Fischer's table with 11-G Mammotome vacuum probes. Imprint samples were examined (Diff-Quick stain, modified Papanicolaou stain and May-Grünwald–Giemsa). The cores were dipped into a CytoRich Red Collection fluid for a few seconds in order to obtain samples with the use of the specimen wash. After the completion of cytological procedures, the core was prepared for routine pathological study. The pathologist was blind to the preliminary cytological results. The cytological and pathological diagnoses were comparatively evaluated.
Results:  The sensitivity of the cytological imprints for cancer was 90%. The specificity of the method for cancer diagnosis was 100%. Two precursor lesions were present in the material: one case of atypical ductal hyperplasia, which was successfully detected, and one case of lobular neoplasia, which escaped detection. The cytological imprints were inadequate in four out of 72 cases (5.6%), but none of them were included within the malignant subgroup.
Conclusions:  Imprint cytology seems to be an important adjunctive tool in the management of patients with non-palpable mammographic solid lesions. Its very satisfactory sensitivity and optimal specificity could establish its use in general clinical practice.     

Fungal Chitin Induces Trained Immunity in Human Monocytes during Cross-talk of the Host with Saccharomyces cerevisiae

The immune system is essential to maintain the mutualistic homeostatic interaction between the host and its micro- and mycobiota. Living as a commensal, Saccharomyces cerevisiae could potentially shape the immune response in a significant way. We observed that S. cerevisiae cells induce trained immunity in monocytes in a strain-dependent manner through enhanced TNFα and IL-6 production upon secondary stimulation with TLR ligands, as well as bacterial and fungal commensals. Differential chitin content accounts for the differences in training properties observed among strains, driving induction of trained immunity by increasing cytokine production and direct antimicrobial activity both in vitro and in vivo. These chitin-induced protective properties are intimately associated with its internalization, identifying a critical role of phagosome acidification to facilitate microbial digestion. This study reveals how commensal and passenger microorganisms could be important in promoting health and preventing mucosal diseases by modulating host defense toward pathogens and thus influencing the host microbiota-immune system interactions.     

The effects of polymorphisms in the DGAT1, leptin and growth hormone receptor gene loci on body energy, blood metabolic and reproductive traits of Holstein cows

The objective of this study was to examine the impact of polymorphisms in the acyl-CoA:diacylglycerol acyltransferase ( DGAT1 ), leptin and growth hormone receptor genes on body energy (body condition score, total body energy content and cumulative effective energy balance) and blood metabolic traits (levels of β-hydroxybutyrate, glucose and non-esterified fatty acids), measured once before the first calving and then repeatedly throughout first lactation in 497 Holstein cows. The influence of the same polymorphisms on cow reproductive performance and health during the first and second lactations was also assessed. Several reproductive traits were considered including interval, conception and insemination traits, as well as incidence of metritis and reproductive problems. Genotyping was performed using PCR-RFLP ( DGAT1 , leptin ) or allele-specific PCR ( growth hormone receptor ). For each locus, the effect of allele substitution on body energy and blood metabolic traits was estimated using random regression models. The same effect on reproductive traits was assessed with single-trait mixed linear models. Significant ( P  <   0.05) effects on specific reproductive traits were observed. DGAT1 and growth hormone receptor alleles responsible for significant increases in milk production were found to have an adverse effect on reproduction, while the leptin allele responsible for significant increase in milk production was linked to marginally increased metritis frequency. Furthermore, the three studied loci were also found to significantly ( P  <   0.05) affect certain body energy and blood metabolic traits. Several associations are published for the first time, but these should be confirmed by other investigators before the polymorphisms are used in gene-assisted selection.     

Vaginal fold prolapse during the last third of pregnancy, followed by normal parturition, in a bitch

This article describes a 1.5-year-old female, Greek Hound dog, weighing 16 kg, presented with a type III vaginal prolapse which occurred during the last third of pregnancy. Trans-abdominal ultrasonography revealed four live foetuses in the uterine horns. The animal was hospitalized and 4 days later gave birth without any interference. Three days later, resection of the prolapsed tissue was performed and the bitch recovered completely. Recurrence of a type I vaginal prolapse was observed 4 months later, during subsequent oestrus. This case is unusual because, although vaginal fold prolapse is mainly seen during proestrus/oestrus or during parturition, it was first noticed 47 days after mating and 13 days before parturition. Furthermore, even though the prolapse of vaginal fold was of type III and of considerable size, parturition proceeded normally. Finally, even though resection of the prolapsed tissue was performed 3 days after parturition, recurrence of vaginal fold oedema (type I) was observed in the subsequent oestrus.     

Morphology of the archaellar motor and associated cytoplasmic cone in Thermococcus kodakaraensis

Archaeal swimming motility is driven by archaella: rotary motors attached to long extracellular filaments. The structure of these motors, and particularly how they are anchored in the absence of a peptidoglycan cell wall, is unknown. Here, we use electron cryotomography to visualize the archaellar basal body in vivo in Thermococcus kodakaraensis KOD1. Compared to the homologous bacterial type IV pilus (T4P), we observe structural similarities as well as several unique features. While the position of the cytoplasmic ATPase appears conserved, it is not braced by linkages that extend upward through the cell envelope as in the T4P, but rather by cytoplasmic components that attach it to a large conical frustum up to 500 nm in diameter at its base. In addition to anchoring the lophotrichous bundle of archaella, the conical frustum associates with chemosensory arrays and ribosome‐excluding material and may function as a polar organizing center for the coccoid cells.