The Concentration of Zn, Fe, Mn, Cu and Se in Fiber Fractions of Legumes in Indonesia

This study was carried out to evaluate concentration of micro minerals (Zn, Fe, Mn, Cu and Se) of forages and their distribution in fiber fraction (neutral detergent fiber/NDF and acid detergent fiber/ADF) in West Sumatra during dry and rainy seasons. Four species of common legume namely Leucaena leucocephala, Centrocema pubescens, Calopogonium mucunoides and Acacia mangium were collected at native pasture during rainy and dry seasons. The results showed that micro minerals concentration of forages and their distribution in fiber fraction varied among species and season. In general, concentration of micro minerals was slightly higher in rainy season compared to dry season either in legumes forages. Data on legume forages showed that 75% of legumes were deficient in Zn and Mn, 62.5 % deficient in Cu and 50 % deficient in Se. There was no species of legume deficient in Fe. Distribution of micro minerals in NDF and ADF were also significantly affected by species and season and depends on the kinds of element measured. Generally, micro minerals were associated in fiber fractions and it yield much higher during dry season compared to rainy season. Iron (Fe) and selenium (Se) in forages were the highest elements associated in NDF and ADF, while the lowest was found in Copper (Cu). (Animal Production 12(2): 105-110 (2010

Additional file 3: of In vitro generation of Sertoli-like and haploid spermatid-like cells from human umbilical cord perivascular cells

Supplementary Figure S1–S4 (PPTX 5065 kb

Initial germ cell to somatic cell ratio impacts the efficiency of SSC expansion <i>in vitro</i>

<p>Spermatogonial Stem Cell (SSC) expansion <i>in vitro</i> remains a major challenge in efforts to preserve fertility among pubertal cancer survivor boys. The current study focused on innovative approaches to optimize SSC expansion. Six- to eight-week-old CD-1 murine testicular samples were harvested by mechanical and enzymatic digestion. Cell suspensions were incubated for differential plating (DP). After DP, we established two experiments comparing single vs. repetitive DP (S-DP and R-DP, respectively) until passage 2 (P2) completion. Each experiment included a set of cultures consisting of 5 floating-to-attached cell ratios (5, 10, 15, 20, and 25) and control cultures containing floating cells only. We found similar cell and colony count drops during P0 in both S- and R-DP. During P2, counts increased in S-DP in middle ratios (10, 15, and especially 20) relative to low and high ratios (5 and 25, respectively). Counts dropped extensively in R-DP after passage 2. The superiority of intermediate ratios was demonstrated by enrichment of GFRα1 by qPCR. The optimal ratio of 20 in S-DP contained significantly increased proportions of GFRα1-positive cells (25.8±5.8%) as measured by flow cytometry compared to after DP (1.9±0.7%, <i>p</i><0.0001), as well as positive immunostaining for GFRα1 and UTF1, with rare Sox9-positive cells. This is the first report of the impact of initial floating-to-attached cell ratios on SSC proliferation <i>in vitro</i>.</p> <p><b>Abbreviations</b>: SSC: spermatogonial stem cells; DP: differential plating; NOA: non-obstructive azoospermia; MACS: magnetic-activated cells sorting; FACS: fluorescence-activated cells sorting </p

The elusive MAESTRO gene: Its human reproductive tissue-specific expression pattern

<div><p>The encoded transcript of the Maestro—<i>Male-specific Transcription in the developing Reproductive Organs</i> (MRO) gene exhibits sexual dimorphic expression during murine gonadal development. The gene has no homology to any known gene and its expression pattern, protein function or structure are still unknown. Previously, studying gene expression in human ovarian cumulus cells, we found increased expression of <i>MRO</i> in lean-type Polycystic Ovarian Syndrome (PCOS) subjects, as compared to controls. In this study, we examined the <i>MRO</i> splice variants and protein expression pattern in various human tissues and cells. We found a differential expression pattern of the <i>MRO</i> 5’-UTR region in luteinized granulosa-cumulus cells and in testicular tissues as compared to non-gonadal tissues. Our study also shows a punctate nuclear expression pattern and disperse cytoplasmic expression pattern of the MRO protein in human granulosa-cumulus cells and in testicular germ cells, which was later validated by western blotting. The tentative and unique features of the protein hampered our efforts to gain more insight about this elusive protein. A better understanding of the tissue-specific <i>MRO</i> isoforms expression patterns and the unique structure of the protein may provide important insights into the function of this gene and possibly to the pathophysiology of PCOS.</p></div

<i>MRO</i> expression in human tissues with FL-248.

<p><i>MRO</i> expression in (A) ovarian tissue of a post-menopausal subject with <b>no active folliculogenesis</b> (age 52); (B) adult normal testis; (C) Typical testicular seminoma; (D) Cerebrum (brain) tissue. Abbreviations: GLC–granulosa-lutein cells, TL–theca lutein cells, SC–Sertoli cells, SG–spermatogonia, PS–primary spermatocyte, SP–spermatids, NP–Neuropil, GL–glial cells. Bar ruler, 100um.</p

Clinical parameters of PCOS and control patients.

<p>Clinical parameters of PCOS and control patients.</p

List of PCR primers.

<p>List of PCR primers.</p

Summary of the MRO splice variants detected in GCs, CCs and human tissues by standard PCR and TaqMan qPCR analysis.

<p>(+) indicates positive detection and intensity (++++ being the highest). (-) indicates no product detected.</p

<i>MRO</i> protein expression in human primary follicle.

<p>Immunofluorescence was performed on an ovarian section from a non-PCOS (A) and a PCOS (B) patients using the FL-248 antibody (4ug/ml). Blocking peptide in (C) demonstrated the reduced fluorescence signal in the tissue. <i>MRO</i> protein is present in GCs and in some stromal cells. <i>MRO</i> expression is indicated in green (Alexa-fluor 488) and nucleus in blue (DAPI). Magnification: x4 and x100; Duration of exposure in 40x: 3886 and 100x: 2000 milliseconds.</p

MRO is detected in both the cytoplasm and nucleus by western Immunoblot.

<p>A) Nuclear and cytoplasmic extracts were prepared from purified GCs, electrophoresed through a 4–12% reducing SDS-PAGE gel, transferred to 0.2um nitrocellulose, blocked, and probed with the MRO FL-248 (Santa Cruz), anti-HDAC1 (Abcam) and anti-Actin (Sigma-Aldrich) antibodies. MRO (~28kDa) is found at similar abundance in both the cytoplasmic and nuclear fraction (top panel). HDAC1 served as a nuclear specific control to ensure the purity of the extract (middle panel) and Actin served as a loading control (bottom panel). B) Protein extracts from portions of GCs from the different isolation method were resolved. GCs pre-wash (Load), after the 1^st wash (post-wash) and after the Ficoll gradient (Post-Ficoll) were probes with the lymphocyte marker anti ἀCD45. Actin served as a loading control (bottom panel).</p