Sven Bocklandt

Epigenetic Predictor of Age

From the moment of conception, we begin to age. A decay of cellular structures, gene regulation, and DNA sequence ages cells and organisms. DNA methylation patterns change with increasing age and contribute to age related disease. Here we identify 88 sites in or near 80 genes for which the degree of cytosine methylation is significantly correlated with age in saliva of 34 male identical twin pairs between 21 and 55 years of age. Furthermore, we validated sites in the promoters of three genes and replicated our results in a general population sample of 31 males and 29 females between 18 and 70 years of age. The methylation of three sites—in the promoters of the EDARADD, TOM1L1, and NPTX2 genes—is linear with age over a range of five decades. Using just two cytosines from these loci, we built a regression model that explained 73% of the variance in age, and is able to predict the age of an individual with an average accuracy of 5.2 years. In forensic science, such a model could estimate the age of a person, based on a biological sample alone. Furthermore, a measurement of relevant sites in the genome could be a tool in routine medical screening to predict the risk of age-related diseases and to tailor interventions based on the epigenetic bio-age instead of the chronological age.

Activation of latent HIV-1 expression by the potent anti-tumor promoter 12-deoxyphorbol 13-phenylacetate

Agents that induce HIV-1 out of latency would be useful adjuvants for currently available anti-retroviral therapy. We report that nanomolar concentrations of 12-deoxyphorbol 13-phenylacetate (DPP), an anti-tumor-promoting phorbol ester originally isolated from a West African plant, induce the expression of HIV-1 in latently infected T cells and render them sensitive to killing by an immunotoxin targeted to the viral envelope glycoprotein. DPP also regulates an extensive series of genes under the control of protein kinase C, including several involved in T cell activation and cytoskeleton reorganization, and represses expression of the HIV-1 receptor CD4 and coreceptor CXCR4. DPP is 20-40-fold more potent than the related phorbol ester prostratin, probably due to its more lipophilic side chain structure. The combination of high potency and anti-tumor promoting activity make DPP an attractive candidate for the adjunctive therapy of persistent HIV-1 infection.

Sex differences in brain and behavior: hormones versus genes.

Sex determination is the commitment of an organism toward male or female development. Traditionally, in mammals, sex determination is considered equivalent to gonadal determination. Since the presence or the absence of the testes ultimately determines the phenotype of the external genitalia, sex determination is typically seen as equivalent to testis determination. But what exactly does sex determine? The endpoint of sex determination is almost invariably seen as the reproductive structures, which represent the most obvious phenotypic difference between the sexes. One could argue that the most striking differences between males and females are not the anatomy of the genitals, but the size of the gametes-considerably larger in females than males. In fact, there could be many different endpoints to sex determination, leading to differences between the sexes: brain sexual differences, behavioral differences, and susceptibility to disease. The central dogma of sexual differentiation, stemming initially from the gonad-transfer experiments of Alfred Jost, is that sexual dimorphisms of all somatic tissues are dependent on the testicular secretion of the developing fetus. In this chapter, we will take the example of sex differences in brain and behavior as an endpoint of sex determination. We will argue that genetic factors play a role in sexually dimorphic traits such as the number of dopaminergic cells in the mesencephalon, aggression, and sexual orientation, independently from gonadal hormones.

Extreme skewing of X chromosome inactivation in mothers of homosexual men

Human sexual preference is a sexually dimorphic trait with a substantial genetic component. Linkage of male sexual orientation to markers on the X chromosome has been reported in some families. Here, we measured X chromosome inactivation ratios in 97 mothers of homosexual men and 103 age-matched control women without gay sons. The number of women with extreme skewing of X-inactivation was significantly higher in mothers of gay men (13/97=13%) compared to controls (4/103=4%) and increased in mothers with two or more gay sons (10/44=23%). Our findings support a role for the X chromosome in regulating sexual orientation in a subgroup of gay men.

A Candidate Gene Study of CYP19 (Aromatase) and Male Sexual Orientation

Aromatase cytochrome P450 (CYP19), which is necessary for the conversion of androgens to estrogens, plays an important role in the sexual differentiation of the brain. To investigate whether differences in the gene encoding the aromatase enzyme influence sexual orientation in men, we conducted linkage, association, and expression analyses in a large sample of homosexual brothers using microsatellite markers in and around CYP19. No linkage was detected, and a gene-specific relative risk of 1.5-fold could be excluded at a lod score of −2. Results of the TDT demonstrated no preferential transmission of any of the CYP19 alleles in this sample. Expression of aromatase mRNA by microarray analysis was not significantly different between heterosexual and homosexual men. These results suggest that variation in the gene for this subunit of the aromatase enzyme complex is not likely to be a major factor in the development of individual differences in male sexual orientation.

Rational Design of Drugs That Induce Human Immunodeficiency Virus Replication

ABSTRACT Drugs that induce human immunodeficiency virus type 1 (HIV-1) replication could be used in combination with highly active antiretroviral therapy (HAART) to reduce the size of the latent reservoir that is in part responsible for viral persistence. Protein kinase C (PKC) is a logical target for such drugs because it activates HIV-1 transcription through multiple mechanisms. Here we show that HIV-1 gene expression can be induced by potent synthetic analogues of the lipid second messenger diacylglycerol (DAG) synthesized on a five-member ring platform that reduces the entropy of binding relative to that of the more flexible DAG template. By varying the alkyl side chains of these synthetic DAG lactones, it was possible to maximize their potency and ability to render latently infected T cells sensitive to killing by an anti-HIV-1 immunotoxin while minimizing the side effects of CD4 and CXCR4 downregulation and tumor necrosis factor alpha upregulation. The two lead compounds, LMC03 and LMC07, regulated a series of PKC-sensitive genes involved in T-cell activation and induced viral gene expression in peripheral blood mononuclear cells from HIV-1-infected individuals. These studies demonstrate the potential for the rational design of agents that, in conjunction with HAART and HIV-specific toxins, can be used to decrease or eliminate the pool of latently infected reservoirs by forcing viral expression.

Prolactin expression in the sheep brain.

Accumulating evidence in rodents suggests that a prolactin locally synthesized and released within the brain can act together with that taken up from the circulation to modulate neuroendocrine responses. The present study was designed to identify the regional patterns of prolactin expression in the adult and developing sheep brain. Specifically, we tested the hypothesis that prolactin is expressed in regions of the adult and fetal sheep brain that are critical in the development of neuroendocrine homeostatic and behavioral functions. The expression of prolactin protein in sheep brain was demonstrated by Western blot analysis and brain prolactin mRNA was detected and sequenced using RT-PCR. In situ hybridization histochemistry revealed that prolactin mRNA was expressed in the medial preoptic area, periventricular preoptic nucleus, bed nucleus of the stria terminalis, and in the paraventricular nucleus of the hypothalamus, particularly the ventral region. The neuroanatomical distribution of prolactin mRNA was best visualized in the fetus and prolactin-immunoreactive neurons could also be identified in late gestation fetuses. Brain prolactin mRNA was expressed as early as day 60 of gestation and increased as the fetus aged and peaked at day 135 (term = 147 days). Prolactin mRNA expression did not exhibit a sex difference in the preoptic area, but in the amygdala prolactin mRNA was significantly higher in females than in males at day 100 of gestation. We conclude that prolactin expressed in adult and fetal sheep brain could be involved in neurodevelopment and/or modulation of the neuroendocrine stress axis, although it is too early to rule out other possibilities given the diverse actions that have been attributed to prolactin.

The Relationship Between Help-Seeking Attitudes and Masculine Norms Among Monozygotic Male Twins Discordant for Sexual Orientation

OBJECTIVE In general, heterosexual men are less favorable to asking for help compared to women and gay men. This can be problematic if a man avoids professional help when he is experiencing significant psychological distress. Yet, it is unclear to what degree such attitudes among men are due to innate differences or social environments. Studying twins provides one avenue for teasing apart these relationships. METHOD We recruited 38 pairs of monozygotic male twins (Mage = 35.87 years, SD = 9.52) raised together and who were discordant for sexual orientation. They completed measures of psychological distress (Symptom Checklist-90-Revised), positive attitudes toward psychological help-seeking behavior, and emphasis with fulfilling traditional masculine norms. RESULTS Contrary to predictions, the heterosexual twins expressed more symptoms of specific distress-hostility (r = .30), paranoid ideation (r = .26), and psychoticism (r = .24)-than their gay cotwins. As predicted, heterosexual men were less favorable to seeking help (r = .25) and expressed greater emphasis on masculine norms (r = .26) than their cotwins. Within each group of men, unique aspects of masculine norms were significantly related to attitudes toward psychological help-seeking behavior. CONCLUSION The findings lend credence to the hypothesis that social environments influence attitudes and behaviors that are stereotypically masculine and potentially detrimental to mens health.

The Biology of Sexual Orientation and Gender Identity

The two largest sex differences in behavior between men and women are sexual orientation and gender identity. Sexual orientation refers to the direction of sexual attraction to the same or other sex/gender. Gender identity refers to peoples inner sense of maleness or femaleness. Although these phenomena are a fundamental part of human life, their underlying mechanisms remain poorly understood. The dominant theory is that prenatal hormones of gonadal origin play a major role in both sexual orientation and gender identity. However, a review of the evidence shows that hormones fail to explain either of these traits in the vast majority of humans. Furthermore, correlations with several parameters, including digit length ratio, handedness, fraternal birth order, and brain anatomy, do not provide clear causal etiology to masculine or feminine sexual behavior. Emerging studies on twins and the application of linkage analysis provide arguments for a genetic influence on sexual orientation and is reviewed in this chapter.