Autism with intellectual disability is associated with increased levels of maternal cytokines and chemokines during gestation.

Immune abnormalities have been described in some individuals with autism spectrum disorders (ASDs) as well as their family members. However, few studies have directly investigated the role of prenatal cytokine and chemokine profiles on neurodevelopmental outcomes in humans. In the current study, we characterized mid-gestational serum profiles of 22 cytokines and chemokines in mothers of children with ASD (N=415), developmental delay (DD) without ASD (N=188), and general population (GP) controls (N=428) using a bead-based multiplex technology. The ASD group was further divided into those with intellectual disabilities (developmental/cognitive and adaptive composite score<70) (ASD+ID, N=184) and those without (composite score⩾70) (ASD-noID, N=201). Levels of cytokines and chemokines were compared between groups using multivariate logistic regression analyses, adjusting for maternal age, ethnicity, birth country and weight, as well as infant gender, birth year and birth month. Mothers of children with ASD+ID had significantly elevated mid-gestational levels of numerous cytokines and chemokines, such as granulocyte macrophage colony-stimulating factor, interferon-γ, interleukin-1α (IL-1α) and IL-6, compared with mothers of children with either ASD-noID, those with DD, or GP controls. Conversely, mothers of children with either ASD-noID or with DD had significantly lower levels of the chemokines IL-8 and monocyte chemotactic protein-1 compared with mothers of GP controls. This observed immunologic distinction between mothers of children with ASD+ID from mothers of children with ASD-noID or DD suggests that the intellectual disability associated with ASD might be etiologically distinct from DD without ASD. These findings contribute to the ongoing efforts toward identification of early biological markers specific to subphenotypes of ASD.

Increased female autosomal burden of rare copy number variants in human populations and in autism families.

Autosomal genetic variation is presumed equivalent in males and females and makes a major contribution to disease risk. We set out to identify whether maternal copy number variants (CNVs) contribute to autism spectrum disorders (ASDs). Surprisingly, we observed a higher autosomal burden of large, rare CNVs in females in the population, reflected in, but not unique to, ASD families. Meta-analysis across control data sets confirms female excess in CNV number (P=2.1 × 10(-5)) and gene content (P=4.1 × 10(-3)). We additionally observed CNV enrichment in ASD mothers compared with control mothers (P=0.03). We speculate that tolerance for CNV burden contributes to decreased female fetal loss in the population and that ASD-specific maternal CNV burden may contribute to high sibling recurrence. These data emphasize the need for study of familial CNV risk factors in ASDs and the requirement of sex-matched comparisons.

Effects of contraceptive use on bone biochemical markers in young women.

The purpose of this study was to compare biochemical markers of bone resorption and formation in young women using different hormonal contraceptive methods. Women aged 18-39 yr who were using depot medroxyprogesterone acetate (DMPA) contraception were recruited for the study; comparison women were matched by age and clinic location. There were 116 women using DMPA, 39 using oral contraceptives containing estrogen and progestin, and 72 not currently using hormonal contraceptives. Biochemical measurements were serum calcium, PTH and osteocalcin, and urine N-telopeptide. Bone density was measured using dual-energy x-ray absorptiometry. The N-telopeptide levels, adjusted for age and other risk factors, were 42.4 +/- 2.3 nmol/mmol creatinine in the DMPA group, 26.2 +/- 3.3 nmol/mmol in the oral contraceptive group, and 35.4 +/- 2.9 nmol/mmol in the nonusers; significant differences were seen in all pairwise comparisons. Osteocalcin levels showed the same pattern, although the difference between the DMPA users and nonusers was not statistically significant. There were no differences among groups in the PTH levels. The bone density at the spine was 1.086 +/- 0.085 g/cm(2) in the DMPA group, 1.103 +/- 0.095 g/cm(2) in the oral contraceptive group, and 1.093 +/- 0.090 g/cm(2) in nonusers (P = 0.051). The results suggest that in women using DMPA bone resorption exceeded bone formation.

Cost comparison of mastectomy versus breast-conserving therapy for early-stage breast cancer.

BACKGROUND: Choice of treatment for early-stage breast cancer depends on many factors, including the size and stage of the cancer, the woman's age, comorbid conditions, and perhaps the costs of treatment. We compared the costs of all medical care for women with early-stage breast cancer cases treated by breast-conserving therapy (BCT) or mastectomy. METHODS: A total of 1675 women 35 years old or older with incident early-stage breast cancer were identified in a large regional nonprofit health maintenance organization in the period 1990 through 1997. The women were treated with mastectomy only (n = 183), mastectomy with adjuvant hormonal therapy or chemotherapy (n = 417), BCT with radiation therapy (n = 405), or BCT with radiation therapy and adjuvant hormonal therapy or chemotherapy (n = 670). The costs of all medical care for the period 1990 through 1998 were computed for each woman, and monthly costs were analyzed by treatment, adjusting for age and cancer stage. All statistical tests were two-sided. RESULTS: At 6 months after diagnosis, the mean total medical care costs for the four groups differed statistically significantly (P:<.001), with BCT being more expensive than mastectomy. The adjusted mean costs were $12 987, $14 309, $14 963, and $15 779 for mastectomy alone, mastectomy with adjuvant therapy, BCT plus radiation therapy, and BCT plus radiation therapy with adjuvant therapy, respectively. At 1 year, the difference in costs was still statistically significant (P:<.001), but costs were influenced more by the use of adjuvant therapy than by type of surgery. The 1-year adjusted mean costs were $16 704, $18 856, $17 344, and $19 081, respectively, for the four groups. By 5 years, BCT was less expensive than mastectomy (P:<.001), with 5-year adjusted mean costs of $41 930, $45 670, $35 787, and $39 926, respectively. Costs also varied by age, with women under 65 years having higher treatment costs than older women. CONCLUSIONS: BCT may have higher short-term costs but lower long-term costs than mastectomy.

Depressive symptoms in users and non-users of depot medroxyprogesterone acetate.

To evaluate the possible effects of depot medroxyprogesterone acetate (DMPA) injectable contraception on depressive symptoms, we conducted a population-based prospective study with women aged 18-39 years old enrolled at a health maintenance organization. At baseline, 183 women used DMPA and 274 were non-users. Data on depressive symptoms and on factors potentially related to DMPA use and depression were collected by questionnaire at 6-month intervals for up to 3 years. In multivariate longitudinal analysis, we found an increased likelihood of reporting depressive symptoms among continuous DMPA users (OR = 1.44; 95% CI = 1.00-2.07) and discontinuers (OR = 1.60; 95% CI = 1.03-2.48) when compared to non-users. Women who discontinued DMPA use had elevated depressive symptoms prior to discontinuation (OR = 2.30; 95% CI = 1.42-3.70) and immediately following discontinuation (OR = 2.46; 95% CI = 1. 46-4.14), and depressive symptoms subsided at subsequent visits relative to non-users. Our prospective analyses found an association between DMPA use and depressive symptoms but further research is needed to determine whether the relationship is causal.

Lack of neurotrophin-3 results in death of spinal sensory neurons and premature differentiation of their precursors.

To understand mechanisms resulting in the absence of two-thirds of spinal sensory neurons in mice lacking NT-3, we have compared dorsal root ganglia development in normal and mutant embryos. The reduction in neurons, achieved by E13, results from several deficits: first, elevated neuronal apoptosis significantly reduces neuronal numbers; second, elevated neurogenesis between E11 and E12, without changes in rates of precursor proliferation or apoptosis, depletes the precursor pool; consequently, the reduced precursor pool prevents increases in neuronal numbers between E12 and E13, when most neurons are born in normal animals. Although deficits occur before final target innervation, we show that NT-3 is expressed at all stages in regions accessible to these neurons or their axons and is only restricted to final targets after innervation.

Neurotrophin-3 is a survival factor in vivo for early mouse trigeminal neurons.

Mice lacking neurotrophin-3 (NT-3) have been shown previously to be born with severe sensory deficits. This study characterizes the developmental course of this deficit in the trigeminal sensory ganglion, which in NT-3 homozygous mutants contains only 35% of the normal number of neurons at birth. At embryonic day 10.5 (E10.5), normal numbers of neurons, as assessed by expression of neurofilament protein and of total cells, are present in the ganglia of mutant homozygotes. During the next 3 d (E10.5-E13.5), virtually all of the deficit develops, after which mutant animals retain only approximately 30% the normal number of neurons. Quantification of neuronal and neuronal precursor numbers in normal and mutant animals reveals that neurons are specifically depleted in the absence of NT-3. A deficiency in precursor proliferation is only seen after most of the neuronal deficit has developed. Numbers of apoptotic cells in the ganglia of mutant animals are elevated during this same interval, indicating that the neuronal deficit is caused, in large part, by increased cell death of embryonic neurons. To determine sources of NT-3 in the trigeminal system, we examined the expression pattern of beta-galactosidase in mice, in which lacZ has replaced the NT-3 coding exon. E10.5-E11.5 embryos exhibit intense reporter expression throughout the mesenchyme and epithelia of the first branchial arch. Beta-galactosidase expression in E13.5 embryos is largely confined to the oral epithelium and the mesenchyme underlying the skin. Throughout the E10.5-E13.5 interval, the trigeminal ganglion and its targets in the CNS do not express reporter activity. We conclude that NT-3 acts principally as a peripherally derived survival factor for early trigeminal neurons.

Identification of integrin alpha 3 beta 1 as a neuronal thrombospondin receptor mediating neurite outgrowth.

Thrombospondins are a family of extracellular matrix proteins expressed throughout the developing nervous system that promote neurite outgrowth in vitro and help mediate the migration of granule cells across the molecular layer in explants of neonatal cerebellum. The receptors mediating these interactions have not previously been identified. In this study, monoclonal antibodies raised to the integrin alpha 3 beta 1 heterodimer are shown to inhibit neurite outgrowth by rat sympathetic neurons on thrombospondin-1. Alpha 3 beta 1 is found to be expressed on the cell body, neurites, and growth cones of sympathetic neurons in vitro and on sympathetic axons passing through the thrombospondin-rich outer sheath of the superior cervical ganglion in vivo, consistent with its role in mediating axon outgrowth. A receptor-ligand binding assay is used to demonstrate the direct binding of immunopurified alpha 3 beta 1 to thrombospondin-1. These results demonstrate a direct interaction between the integrin alpha 3 beta 1 and thrombospondin-1, which mediates neurite outgrowth in vitro and is likely to mediate the same interactions in vivo.

Expression and in vitro function of beta 1-integrin laminin receptors in the developing avian ciliary ganglion.

In chick development, ciliary ganglion (CG) neurons go through a period of axon extension from approximately embryonic day (E)4 to E8, followed by a period of synaptogenesis and neuronal cell death. By examining the immunohistochemical localization of laminin, in conjunction with Dil labeling of the ciliary nerve projection, we have determined that the pathway taken by these neurons is rich in laminin expression. Therefore, laminins are good candidate molecules for mediating outgrowth of these neurons in vivo. In vitro, the ability of CG neurons to extend neurites on laminin-1 (EHS laminin, alpha 1 beta 1 gamma 1) is maximal up to E8, then declines dramatically. CG neuron outgrowth on laminin-1 requires the activity of beta 1-class integrins. We have used subunit-specific antibodies to determine which of the five beta 1-containing heterodimers known to be laminin receptors (alpha 1 beta 1, alpha 2 beta 1, alpha 6 beta 1, alpha 7 beta 1) are expressed, and which mediate neurite outgrowth. While we could not detect expression of alpha 2 or alpha 7, we have found that alpha 1, alpha 3 beta 1, and alpha 6 beta 1 are expressed on the surface of ciliary ganglion neuron cell bodies and axons, both in vitro and in vivo. Furthermore, antibodies against alpha 3 and alpha 6, but not alpha 1, interfered with CG neurite outgrowth on laminin-1 in vitro. Taken together, these data suggest that interactions of cell surface alpha 3 beta 1 and alpha 6 beta 1 integrins with laminin-1 are likely to mediate growth of CG neurons during pathfinding in vivo.

The cadherin-binding specificities of B-cadherin and LCAM.

The cadherin family of calcium-dependent cell adhesion molecules plays an important part in the organization of cell adhesion and tissue segregation during development. The expression pattern and the binding specificity of each cadherin are of principal importance for its role in morphogenesis. B-Cadherin and LCAM, two chicken cadherins, have similar, but not identical, spatial and temporal patterns of expression. To examine the possibility that they might bind to one another in a heterophilic manner, we generated, by cDNA transfection, L-cell lines that express LCAM or B-cadherin. We then examined the abilities of these cells to coaggregate with each other and with other cadherin-expressing cells in short-term aggregation assays. The B-cadherin- and the LCAM-expressing cell lines segregate from P-, N-, or R-cadherin-expressing cells. B-cadherin- and LCAM-expressing cell lines, however, appear to be completely miscible, forming large mixed aggregates. Chick B-cadherin and murine E-cadherin also form mixed aggregates, indistinguishable from homophilic aggregates. Murine E-cadherin and chick LCAM coaggregate less completely, suggesting that the heterophilic interactions of these two cell lines are weak relative to homophilic interactions. These data suggest that heterophilic interactions between B-cadherin and LCAM are important during avian morphogenesis and help identify the amino acids in the binding domain that determine cadherin specificity.