PGRMC1 participates in late events of bovine granulosa cells mitosis and oocyte meiosis.

Progesterone Receptor Membrane Component 1 (PGRMC1) is expressed in both oocyte and ovarian somatic cells, where it is found in multiple cellular sub-compartments including the mitotic spindle apparatus. PGRMC1 localization in the maturing bovine oocytes mirrors its localization in mitotic cells, suggesting a possible common action in mitosis and meiosis. To test the hypothesis that altering PGRMC1 activity leads to similar defects in mitosis and meiosis, PGRMC1 function was perturbed in cultured bovine granulosa cells (bGC) and maturing oocytes and the effect on mitotic and meiotic progression assessed. RNA interference-mediated PGRMC1 silencing in bGC significantly reduced cell proliferation, with a concomitant increase in the percentage of cells arrested at G2/M phase, which is consistent with an arrested or prolonged M-phase. This observation was confirmed by time-lapse imaging that revealed defects in late karyokinesis. In agreement with a role during late mitotic events, a direct interaction between PGRMC1 and Aurora Kinase B (AURKB) was observed in the central spindle at of dividing cells. Similarly, treatment with the PGRMC1 inhibitor AG205 or PGRMC1 silencing in the oocyte impaired completion of meiosis I. Specifically the ability of the oocyte to extrude the first polar body was significantly impaired while meiotic figures aberration and chromatin scattering within the ooplasm increased. Finally, analysis of PGRMC1 and AURKB localization in AG205-treated oocytes confirmed an altered localization of both proteins when meiotic errors occur. The present findings demonstrate that PGRMC1 participates in late events of both mammalian mitosis and oocyte meiosis, consistent with PGRMC1's localization at the mid-zone and mid-body of the mitotic and meiotic spindle.

Parotid abscess: mini-pictorial essay.

Bacterial and viral infection of the intraparotideal and periparotideal lymph nodes, or of the glandular parenchyma, can lead to inflammatory and subsequently suppurative changes, which can result in abscess formation. Sonography allows a detailed morphological evaluation of the gland and has an important value, complementary to clinical examination, in the study of parotid inflammatory diseases. Specifically, sonography defines lesion's characteristics and, in a large number of cases, also its nature and, among the various imaging techniques, it is considered first-line imaging for the evaluation of parotid abscesses, which are characterized by hypo-anechoic lesions, with irregular margins. Color Doppler signals are peripheral to the abscess. If combined with color Doppler, sonography is important not only in diagnosis, but also in the evaluation of therapeutic efficacy and as a guide to the aspiration and drainage of abscesses. The rarity of some of these lesions led us to publish this mini-pictorial essay.

Use of ultrasound-magnetic resonance image fusion to guide sacroiliac joint injections: a preliminary assessment.

BACKGROUND: The sacroiliac joint is one of the sources of chronic lower back pain. Intra-articular injections of anesthetic drugs and/or steroids are currently used in these cases for diagnostic and therapeutic purposes. However, given the anatomic and functional complexity of the joint, imaging guidance is mandatory during such procedures. In this context, the technique of fusing images obtained with two different modalities can often overcome the limitations and enhance the advantages of single-modality guidance. AIM: The aim of this study was to evaluate the technique of ultrasound (US)-magnetic resonance (MR) image fusion to guide intra-articular injections of drugs into the sacroiliac joint. MATERIALS AND METHODS: We evaluated seven sacroiliac joints in six patients with sacroiliac pain syndrome (four females, two males; mean age 59 years; range 46-76 years). Five were candidates for radiofrequency thermolysis, and a therapeutic nerve block was performed in the sixth. Using the volume navigation system, we fused three-dimensional MR images with simultaneously acquired real-time ultrasound images and used them to guide the intra-articular injections. RESULTS: In all patients, spatial accuracy was considered excellent, with definition of registration errors of less than 3 mm. The diagnostic blocks produced positive results in all patients with 80 % reductions in pain (measured with a Numerical Rating Scale, NRS) relative to baseline. The patient who underwent the therapeutic nerve block experienced complete resolution of symptoms that has been maintained over time. There were no complications. CONCLUSIONS: US-MR imaging fusion guidance of sacroiliac joint injections is feasible and effective, in accordance with the data in the literature. The use of the MR for three-dimensional imaging eliminates the risk of radiation exposure.

The genomic and proteomic blueprint of mouse megakaryocytes derived from embryonic stem cells.

BACKGROUND: Platelets are specialized cells, produced by megakaryocytes (MKs) in the bone marrow, which represent the first defense against hemorrhage. There are many diseases where platelet production or function is impaired, with severe consequences for patients. Therefore, new insights into the process of MK differentiation and platelet formation would have a major impact on both basic and clinical research. OBJECTIVES: Embryonic stem (ES) cells represent a good in vitro model to study the differentiation of MKs, with the possibility of being genetically engineered and constituting an unlimited source of MKs. However, lack of knowledge about the molecular identity of ES-derived MKs (ES-MKs) may prevent any further development and application of this model. METHODS: This paper presents the first comprehensive transcriptional and proteome profile analyses of mouse ES-MKs in comparison with MKs derived from mouse fetal liver progenitors (FL-MKs). RESULTS: In ES-MKs we found a down-regulation of cytoskeleton proteins, specific transcription factors and membrane receptors at both transcriptional and protein levels. At the phenotypic level, this molecular blueprint was displayed by ES-MKs' lower polyploidy, lower nuclear/cytoplasm ratio and reduced capacity to form proplatelets and releasing platelets. CONCLUSIONS: Overall our data demonstrate that ES-MKs represent a useful model to clarify many aspects of both MK physiology and pathological conditions where impaired MK functions are related to defective MK development, as in inherited thrombocytopenias and primary myelofibrosis.

Single-cell quantitative RT-PCR analysis of Cpt1b and Cpt2 gene expression in mouse antral oocytes and in preimplantation embryos.

Fatty acids represent an important energy source for preimplantation embryos. Fatty acids oxidation is correlated with the embryo oxygen consumption which remains relatively constant up to the 8-cell stage, but suddenly increases between the 8-cell and morula stages. The degradation of fatty acids occurs in mitochondria and is catalyzed by several carnitine acyl transferases, including two carnitine palmitoyl transferases, CPT-I and CPT-II. We have carried out a study to determine the relative number of transcripts of Cpt1b and Cpt2 genes encoding for m-CPT-I and CPT-II enzymes, during mouse preimplantation development. Here we show that Cpt1b transcripts are first and temporally detected at the 2-cell stage and reappear at the morula and blastocyst stage. Cpt2 transcripts decrease following fertilization to undetectable levels and are present again later at the morula stage. These results show that transcription of both Cpt1b and Cpt2 is triggered at the morula stage, concomitantly with known increasing profiles of oxygen uptake and fatty acids oxidation. Based on the number of Cpt2 transcripts detected, we could discriminate the presence of two groups of embryos with high and low number of transcripts, from the zygote throughout preimplantation development. To further investigate if the establishment of these two groups of embryos occurs prior to fertilization, we have analyzed the relative number of transcripts of both genes in antral and ovulated MII oocytes. As for preimplantation embryos, MII oocytes show two groups of Cpt2 expression. Antral oocytes, classified according to their chromatin configuration in SN (surrounded nucleolus, in which the nucleolus is surrounded by a rim of Hoechst-positive chromatin) and NSN (not surrounded nucleolus, in which this rim is absent), show three groups with different numbers of Cpt2 transcripts. All NSN oocytes have a number of Cpt2 transcripts doubled compared to that of the group of MII oocytes with high expression. Instead, SN oocytes could be singled out into two groups with high and low numbers of Cpt2 transcripts, similar to those found in MII oocytes. The results of this study point out a correlation between the timing of fatty acids oxidation during preimplantation development and the expression of two genes encoding two enzymes involved in the oxidative pathway. Furthermore, although the biological meaning for the presence of two groups of oocytes/embryos with different levels of Cpt2 transcripts remains unclear, the data obtained suggest a possible correlation between the levels of Cpt2 expression and embryo developmental competence.

Meiotic recombination and spermatogenic impairment in Mus musculus domesticus carrying multiple simple Robertsonian translocations.

We quantitatively analyzed the spermatogenic process, including evaluation of seminiferous tubules with defective cycles, rates of germ cell death and sperm morphology, in adult male mice with standard telocentric chromosomes (2n = 40, CD1 strain), homozygous (2n = 24, Mil II population) and heterozygous (2n = 24 x 40) for Robertsonian (Rb) rearrangements. The animals were analyzed at three different ages: three, five and seven months after birth. The number and position of crossover events were also determined by chiasmata counting and immunostaining with an antibody against mouse MLH1 protein. Our analysis of spermatogenesis confirms the impairment of the spermatogenic process in multiple simple heterozygotes due to both germ cell and abnormal sperm morphology. The detrimental effects exerted by Rb heterozygosities were found to be at least partially buffered with time: the frequency of defective tubules was lower and germ cell survival and sperm morphology better in 7-month-old animals than in the 3- and 5-month-old mice. While there are previously published data on germ cell death in multiple simple heterozygotes, this is the first report of a partial rescue of spermatogenesis with time. The mean frequency of MLH1 foci was lower in Rb homozygous and heterozygous mice than in mice carrying all telocentric chromosomes. The lower number of foci in Rb mice can be ascribed to a decrease in the number of multiple chiasmata and the maintenance of single chiasmata preferentially located in the terminal region of both the telocentric and metacentric chromosomes.