Ex vivo Expansion and Differentiation of Mesenchymal Stem Cells from Goat Bone Marrow

Objective(s)Mesenchymal stem cells (MSCs) from large animals as goat which is genetically more closely related tohuman have rarely been gained attentions. The present study tried to isolate and characterize MSCs fromgoat bone marrow.Materials and MethodsFibroblastic cells appeared in goat marrow cell culture were expanded through several subcultures.Passaged-3 cells were then differentiated among the osteogenic, adipogenic and chondrogenic cell lineagesto determine their MSC nature. Differentiations were determined by RT-PCR analysis of related geneexpression. To identify the best culture conditions for propagation, passage-3 cells were plated either atvarying cell densities or different fetal bovine serum (FBS) concentrations for a week, at the end of whichthe cultures were statistically compared with respect to the cell proliferation. In this study, we alsodetermined goat MSC population doubling time (PDT) as the index of their in vitro expansion rate.ResultsPassage-3 fibroblastic cells tended to differentiate into skeletal cell lineages. This was evident in bothspecific staining as well as the specific gene expression profile. Moreover, there appeared to be moreexpansion when the cultures were initiated at 100 cells/cm2 in a medium supplemented with 15% FBS. Arelatively short PDT (24.94±2.67 hr) was a reflection of the goat MSC rapid rate of expansion.ConclusionTaken together, fibroblastic cells developed at goat marrow cell culture are able to differentiate into skeletalcell lineages. They undergo extensive proliferation when being plated at low cell density in 15% FBSconcentration.Keywords: Adipogenesis, Bovine serum, Cell seeding density, Chondrogenesis, Goat mesenchymal stemcells, Osteogenesi

Early Determinants of Childhood Blood Pressure at the Age of 6 Years:The GECKO Drenthe and ABCD Study Birth Cohorts

Background There is still uncertainty about the nature and relative impact of early determinants on childhood blood pressure. This study explored determinants of blood pressure at the age of 6 years in 2 Dutch birth cohorts. Methods and Results Results of hierarchical multiple linear regression analyses in GECKO (Groningen Expert Center for Kids With Obesity) Drenthe study (n=1613) were replicated in ABCD (Amsterdam Born Children and Their Development) study (n=2052). All analyses were adjusted for child's age, sex, height, and body mass index (BMI), and maternal education and subsequently performed in the combined sample. No associations were found between maternal smoking during pregnancy and childhood blood pressure. In the total sample, maternal prepregnancy BMI was positively associated with systolic blood pressure (SBP) (β [95% CI], 0.09 [0.02-0.16] mm Hg) and diastolic blood pressure (β [95% CI], 0.11 [0.04-0.17] mm Hg). Children of women with hypertension had higher SBP (β [95% CI], 0.98 [0.17-1.79] mm Hg). Birth weight standardized for gestational age was inversely associated with SBP (β [95% CI], -6.93 [-9.25 to -4.61] mm Hg) and diastolic blood pressure (β [95% CI], -3.65 [-5.70 to -1.61] mm Hg). Longer gestational age was associated with lower SBP (β [95% CI] per week, -0.25 [-0.42 to -0.08] mm Hg). Breastfeeding for 1 to 3 months was associated with lower SBP (β [95% CI], -0.96 [-1.82 to -0.09] mm Hg) compared with no or <1 month of breastfeeding. Early BMI gain from the age of 2 to 6 years was positively associated with SBP (β [95% CI], 0.41 [0.08-0.74] mm Hg) and diastolic blood pressure (β [95% CI], 0.37 [0.07-0.66] mm Hg), but no effect modification by birth weight was found. Conclusions Higher maternal prepregnancy BMI, maternal hypertension, a relatively lower birth weight for gestational age, shorter gestational age, limited duration of breastfeeding, and more rapid early BMI gain contribute to higher childhood blood pressure at the age of 6 years

Genome-wide analysis identifies 12 loci influencing human reproductive behavior.

The genetic architecture of human reproductive behavior-age at first birth (AFB) and number of children ever born (NEB)-has a strong relationship with fitness, human development, infertility and risk of neuropsychiatric disorders. However, very few genetic loci have been identified, and the underlying mechanisms of AFB and NEB are poorly understood. We report a large genome-wide association study of both sexes including 251,151 individuals for AFB and 343,072 individuals for NEB. We identified 12 independent loci that are significantly associated with AFB and/or NEB in a SNP-based genome-wide association study and 4 additional loci associated in a gene-based effort. These loci harbor genes that are likely to have a role, either directly or by affecting non-local gene expression, in human reproduction and infertility, thereby increasing understanding of these complex traits

Epigenetic editing as a novel approach to modulate expression of key genes in cancer

In dit proefschrift wordt eerst beschreven welke epigenetische veranderingen er in borstkankercellen gevonden zijn. Vervolgens is een nieuwe techniek ontwikkeld die dergelijke epigenetische mutaties kan herstellen: Epigenetische Editing. Een Epigenetische Editor bestaat uit een DNA bindend domain gekoppeld aan een enzymatisch domain, welke epigenetische markeringen kan aanbrengen of verwijderen. Het DNA bindend domain wordt zo gemaakt dat het aan 18 baseparen in het DNA bindt, deze combinatie van 18 baseparen geeft een unieke adressering in het hele menselijke DNA. De genen die in dit proefschrift verder onderzocht worden zijn overactieve oncogenen (Her2/neu, estrogen receptor ESR1) en een epigenetisch geïnactiveerd tumor suppressor gene. Met behulp van Epigenetische Editing konden remmende epigenetische veranderingen worden aangebracht die inderdaad de genexpressie remden. Deze remming resulteerde voor zowel Her2/neu als ESR1 in een verminderde kankercelgroei. EPB41L3, het tumor suppressor gen, komt niet tot expressie in verschillende typen kanker. Wij konden EPB41L3 tot expressie brengen door middel van zogenaamde Artificiële Transcriptie Factoren (bestaande uit een DNA bindend domein gefuseerd aan een transcriptionele activator) in kankercellen en dit resulteerde inderdaad in celdood. In combinatie met andere epigenetische geneesmiddelen, kon de duur van de expressie van dit epigenetisch inactieve gen worden verlengd. Verstoorde epigenetische mechanismen zijn nu gevonden in veel ziekten, ook in kanker. Chemische remmers van epigenetische enzymen zijn goedgekeurd als behandeling voor kanker. Deze middelen werken echter op alle genen in het DNA, met mogelijke bijwerkingen. Epigenetische Editing is een innovatieve benadering om blijvend op specifieke genen effect te hebben. This thesis is about developing a targeted approach to write or erase epigenetic modifications onto a gene of interest (Epigenetic Editing) to modulate its expression. An Epigenetic Editing tool consists of a DNA binding domain (DBD) linked to a catalytic domain of epigenetic enzymes. Using the Epigenetic Editing, inactive epigenetic modifications were written on overexpressed genes in cancer, which were associated with their downregulation and cell growth inhibition. A tumor suppressor gene was upregulated using an artificial transcription factor (composed of a DBD fused to a transcription activator domain) in cancer cells and apoptosis was induced in the treated cells. Since aberrant epigenetic modifications are associated with cancer, the role of epigenetics in breast cancer and the potential epigenetic-therapies are intensively investigated; this is also reviewed in this thesis.