Role of synovial lymphatic function in osteoarthritis.

BACKGROUND: Osteoarthritis (OA) affects the entire joint, initially with a low degree of inflammation. Synovitis is correlated with the severity of OA clinical symptoms and cartilage degradation. The synovial lymphatic system (SLS) plays a prominent role in clearing macromolecules within the joint, including the pro-inflammatory cytokines in arthritic status. Scattered evidence shows that impaired SLS drainage function leads to the accumulation of inflammatory factors in the joint and aggravates the progression of OA, and the role of SLS function in OA is less studied. DESIGN: This review summarizes the current understanding of synovial lymphatic function in OA progression and potential regulatory pathways and aims to provide a framework of knowledge for the development of OA treatments targeting lymphatic structure and functions. RESULTS: SLS locates in the subintima layer of the synovium and consists of lymphatic capillaries and lymphatic collecting vessels. Vascular endothelial growth factor C (VEGF-C) is the most critical regulating factor of lymphatic endothelial cells (LECs) and SLS. Nitric oxide production-induced impairment of lymphatic muscle cells (LMCs) and contractile function may attribute to drainage dysfunction. Preclinical evidence suggests that promoting lymphatic drainage may help restore intra-articular homeostasis to attenuate the progression of OA. CONCLUSION: SLS is actively involved in the homeostatic maintenance of the joint. Understanding the drainage function of the SLS at different stages of OA development is essential for further design of therapies targeting the function of these vessels.

Radula and mandible of Biomphalaria (Planorbidae) of São Paulo, Brazil

The radular apparatus of Biomphalaria occidentalis Paraense, 1981, Biomphalaria oligoza Paraense, 1971, Biomphalaria peregrina (d’Orbigny, 1835), Biomphalaria schrammi (Crosse, 1864) and Biomphalaria tenagophila (d’Orbigny, 1835) from the Metropolitan Region of São Paulo, Brazil were analyzed from Scanning Electron Microscope (SEM) micrographs. The data support the usage of characters of central, lateral and intermediate teeth in the species identification of Biomphalaria. The use of SEM provided additional quantitative and structural details to radula descriptions, which are extremely important for elucidating the taxonomy of Biomphalaria species...

Molecular and morphological identification of Biomphalaria species from the state of São Paulo, Brazil

DNA barcoding and morphological characters were used to identify adult snails belonging to the genus Biomphalaria from 17 municipalities in the state of São Paulo, Brazil. The DNA barcode analysis also included twenty-nine sequences retrieved from GenBank. The final data set of 104 sequences of the mitochondrial cytochrome oxidase I (COI) gene was analyzed for K2P intraspecific and interspecific divergences, through tree-reconstruction methods (Neighbor-Joining, Maximum Likelihood and Bayesianinference), and by applying different models (ABGD, bPTP, GMYC) to partition the sequences according to the pattern of genetic variation. Twenty-seven morphological parameters of internal organs were used to identify specimens. The molecular taxonomy of Biomphalaria agreed with the morphological identification of specimens from the same collection locality. DNA barcoding may therefore be a useful supporting tool for identifying Biomphalaria snails in areas at risk for schistosomiasis...

Infrapatellar fat pad aggravates degeneration of acute traumatized cartilage: a possible role for interleukin-6.

OBJECTIVES: The infrapatellar fat pad (IPFP), which is located underneath the patella, close to cartilage surfaces, functions in distributing mechanical load and has been shown to produce cytokines. This study aims to assess the involvement of the IPFP in the progression of post-traumatic osteoarthritis (OA) through investigating the crosstalk between the IPFP and injured cartilage in vitro. METHODS: A single blunt impact (36 MPa) on healthy bovine articular cartilage explants was used to generate traumatized cartilage. Conditioned media from IPFP and traumatized cartilage (FP-CM and TC-CM) were prepared separately. After culturing in FP-CM, the posttraumatic cartilage explants were analyzed for expression of cartilage degeneration associated genes and secretion of the interleukin (IL)-6, into the culture medium. The effect of traumatized cartilage on IPFP was studied by treating IPFP-derived adipocytes and IPFP adipose-derived stromal cells (ADSC) with TC-CM followed by analysis of cytokine expression. RESULTS: FP-CM aggravated glycosaminoglycan (GAG) release in traumatized cartilage, but did not significantly affect healthy cartilage. FP-CM raised gene expression of cyclooxygenase-2, inducible nitric oxide synthase, and IL-6 in traumatized cartilage explants, and lowered expression of tissue inhibitor of metalloproteinases-1, 2, 3, compared to non-conditioned medium. Of particular significance is that medium IL-6 levels increased substantially in both FP-CM and FP-CM treated traumatized cartilage cultures. Extrinsic IL-6 treatment of traumatized cartilage simulated part of the effects of FP-CM. TC-CM elevated levels of IL-6 expression in IPFP derived adipocytes and ADSCs. CONCLUSIONS: IPFP aggravates post-traumatized cartilage degeneration, and IL-6 is a candidate tissue degeneration mediator.

Experimental study on reproduction of the freshwater snail Biomphalaria tenagophila (d’Orbigny, 1835)

The freshwater snail Biomphalaria tenagophila is one of the most important intermediate hosts of Schistosoma mansoni in the Neotropical region. It is a simultaneous hermaphrodite able to reproduce either by cross- or self-fertilization (SF). In this study, we investigated the mode of reproduction in a laboratory line of the snail seeking to understand the contribution of SF and cross-fertilization (CF) in the natural history of the species. Thirteen mature mutant albino B. tenagophila were paired during 24 h with wild pigmented snails (1alb:1pig). After being paired for 24 h, the albino snails were isolated in glass aquaria and during 70 days the number of albino and pigmented embryos was counted weekly. The excess rather than a deficit of albino snails obtained in the progeny supports the hypothesis that, in B. tenagophila, the mode of reproduction occurs by cross plus SF. The higher rate of selfing rather than CF progeny shows the potential of the species to colonize new, intermittent and unstable freshwater ecosystems...

Laboratório de Bioquímica e Biologia Molecular - LBBM

O Laboratório de Bioquímica e Biologia Molecular (LBBM) se aproxima hoje de um espaço de ciência, de prática e aplicação do conhecimento genético, evolutivo, ecológico, parasitológico, com alto poder de resolução de questões aplicadas à vigilância epidemiológica de doenças. As atividades do LBBM não podem ser resumidas ao simples uso da biologia molecular, como uma técnica ou um instrumento, como por exemplo, um microscópio...

Parâmetros aplicados à análise da produção científica da Superintendência de Controle de Endemias durante os anos 1975 a 2014

As atividades de pesquisa científica na Superintendência de Controle de Endemias (Sucen) respondem à missão institucional de produzir conhecimento para atender às demandas da sociedade quanto ao controle de vetores de doenças transmissíveis. A produção científica merece destaque, pois é através dela que o conhecimento produzido é difundido e democratizado. Desta forma são adquiridas informações para a solução de problemas ou adoção de alternativas para o desenvolvimento e integração sustentável de atividades específicas...

Functional cartilage repair capacity of de-differentiated, chondrocyte- and mesenchymal stem cell-laden hydrogels in vitro.

OBJECTIVE: The long-term performance of cell-seeded matrix-based cartilage constructs depends on (1) the development of sufficient biomechanical properties, and (2) lateral integration with host tissues, both of which require cartilage-specific matrix deposition within the scaffold. In this study, we have examined the potential of tissue-engineered cartilage analogs developed using different cell types, i.e., mesenchymal stem cells (MSCs) vs chondrocytes and de-differentiated chondrocytes, in an established "construct in cartilage ring" model. DESIGN: Cell-laden constructs of differentiated chondrocytes, de-differentiated chondrocytes after two, five or eight population doublings, and MSCs were either implanted into a native cartilage ring immediately after fabrication (immature group) or pre-treated for 21 days in a transforming growth factor-ß3 (TGF-ß3) containing medium prior to implantation. After additional culture for 28 days in a serum-free, chemically defined medium, the extent of lateral integration, and biochemical and biomechanical characteristics of the implants as hybrid constructs were assessed. RESULTS: The quality of integration, the amount of accumulated cartilage-specific matrix components and associated biomechanical properties were found to be highest when using differentiated chondrocytes. De-differentiation of chondrocytes negatively impacted the properties of the implants, as even two population doublings of the chondrocytes in culture significantly lowered cartilage repair capacity. In contrast, MSCs showed chondrogenic differentiation with TGF-ß3 pre-treatment and superior integrational behavior. CONCLUSIONS: Chondrocyte expansion and de-differentiation impaired the cell response, resulting in inferior cartilage repair in vitro. With TGF-ß3 pre-treatment, MSCs were able to undergo sustained chondrogenic differentiation and exhibited superior matrix deposition and integration compared to de-differentiated chondrocytes.

PTHrP overexpression partially inhibits a mechanical strain-induced arthritic phenotype in chondrocytes.

OBJECTIVE: Cell-based tissue engineering strategies are currently in clinical use and continue to be developed at a rapid pace for the repair of cartilage defects. Regardless of the repair methodology, chondrocytes within newly regenerated cartilage remain susceptible to the abnormal inflammatory and mechanical environments that underlie osteoarthritic disease, likely compromising the implant's integration, function, and longevity. The present study investigates the use of parathyroid hormone-related peptide (PTHrP) overexpression for chondroprotection. DESIGN: Bovine articular chondrocytes were transfected with human PTHrP (hPTHrP) constructs (1-141 or 1-173) and subjected to injurious cyclic tensile strain (CTS; 0.5 Hz and 16% elongation) for 48 h. mRNA expression of matrix remodeling, inflammatory signaling, hypertrophic, and apoptotic genes were examined with real-time reverse transcription polymerase chain reaction. Nitric oxide (NO) and prostaglandin E2 (PGE2) production were measured using the Griess assay and enzyme immunoassay (EIA), respectively. RESULTS: CTS-induced an arthritic phenotype in articular chondrocytes as indicated by increased gene expression of collagenases and aggrecanases and increased production of NO and PGE2. Additionally, CTS increased collagen type X (Col10a1) mRNA expression, whereas overexpression of either hPTHrP isoform inhibited CTS-induced Col10a1 gene expression. However, hPTHrP 1-141 augmented CTS-induced NO and PGE2 production, and neither hPTHrP isoform had any significant effect on apoptotic genes. CONCLUSIONS: Our results suggest that chondrocytes overexpressing PTHrP resist mechanical strain-induced hypertrophic-like changes. Therapeutic PTHrP gene transfer may be considered for chondroprotection applications in newly regenerated cartilage.

A nanofibrous cell-seeded hydrogel promotes integration in a cartilage gap model.

The presence of a defect in mature articular cartilage can lead to degenerative changes of the joint. This is in part caused by the inability of cartilage to regenerate tissue that is capable of spanning a fissure or crack. In this study, we hypothesized that introduction of a biodegradable cell-seeded nanofibrous hydrogel, Puramatrix(), into a cartilage gap would facilitate the generation of a mechanically stable interface. The effects of chondrocyte incorporation within the hydrogel and supplementation with transforming growth factor-beta3 (TGFbeta3), a known regulator of cell growth and differentiation, on cartilage integration were examined mechanically and histologically as a function of cell density and incubation time. When supplemented with TGFbeta3, the cell-seeded hydrogel exhibited abundant matrix generation within the hydrogel and a corresponding increase in maximum push-out stress as compared to all other groups. Furthermore, initial cell seeding density affected interfacial strength in a time-dependent manner. This study suggests that a cell-seeded TGFbeta3-supplemented hydrogel can encourage integration between two opposing pieces of articular cartilage.

Beta4 integrin promotes osteosarcoma metastasis and interacts with ezrin.

The development of pulmonary metastasis is the major cause of death in osteosarcoma, and its molecular basis is poorly understood. In this study, we show that beta4 integrin is highly expressed in human osteosarcoma cell lines and tumor samples. Furthermore, highly metastatic MNNG-HOS cells have increased levels of beta4 integrin. Suppression of beta4 integrin expression by shRNA and disruption of beta4 integrin function by transfection of dominant-negative beta4 integrin was sufficient to revert this highly metastatic phenotype in the MNNG-HOS model without significantly affecting primary tumor growth. These findings suggest a role for beta4 integrin expression in the metastatic phenotype in human osteosarcoma cells. In addition, we identified a previously uncharacterized interaction between beta4 integrin and ezrin, a membrane-cytoskeletal linker protein that is implicated in the metastatic behavior of osteosarcoma. The beta4 integrin-ezrin interaction appears to be critical for maintenance of beta4 integrin expression. These data begin to integrate ezrin and beta4 integrin expression into a model of action for the mechanism of osteosarcoma metastases.

Mesenchymal progenitor cells derived from traumatized human muscle.

Mesenchymal stem cells (MSCs) derived from adult tissues are an important candidate cell type for cell-based tissue engineering and regenerative medicine. Currently, clinical applications for MSCs require additional surgical procedures to harvest the autologous MSCs (i.e. from bone marrow) or commercial allogeneic alternatives. We have recently identified a population of mesenchymal progenitor cells (MPCs) in traumatized muscle tissue that has been surgically debrided from traumatic orthopaedic extremity wounds. The purpose of this study was to evaluate whether MPCs derived from traumatized muscle may provide a clinical alternative to bone-marrow MSCs, by comparing their morphology, proliferation capacity, cell surface epitope profile and differentiation capacity. After digesting the muscle tissue with collagenase, the MPCs were enriched by a direct plating technique. The morphology and proliferation rate of the muscle-derived MPCs was similar to bone-marrow derived MSCs. Both populations expressed cell surface markers characteristic for MSCs (CD 73, CD 90 and CD105), and did not express markers typically absent on MSCs (CD14, CD34 and CD45). After 21 days in specific differentiation media, the histological staining and gene expression of the MPCs and MSCs was characteristic for differentiation into osteoblasts, chondrocytes and adipocytes, but not into myoblasts. Our findings demonstrate that traumatized muscle-derived MPCs exhibit a similar phenotype and resemble MSCs derived from the bone marrow. MPCs harvested from traumatized muscle tissue may be considered for applications in tissue engineering and regenerative medicine following orthopaedic trauma requiring circumferential debridement.

The beneficial effect of delayed compressive loading on tissue-engineered cartilage constructs cultured with TGF-beta3.

OBJECTIVE: To determine whether the functional properties of tissue-engineered constructs cultured in a chemically-defined medium supplemented briefly with TGF-beta3 can be enhanced with the application of dynamic deformational loading. METHODS: Primary immature bovine cells (2-3 months old) were encapsulated in agarose hydrogel (2%, 30 x 10(6)cells/ml) and cultured in chemically-defined medium supplemented for the first 2 weeks with transforming growth factor beta 3 (TGF-beta3) (10 microg/ml). Physiologic deformational loading (1 Hz, 3 h/day, 10% unconfined deformation initially and tapering to 2% peak-to-peak deformation by day 42) was applied either concurrent with or after the period of TGF-beta3 supplementation. Mechanical and biochemical properties were evaluated up to day 56. RESULTS: Dynamic deformational loading applied concurrently with TGF-beta3 supplementation yielded significantly lower (-90%) overall mechanical properties when compared to free-swelling controls. In contrast, the same loading protocol applied after the discontinuation of the growth factor resulted in significantly increased (+10%) overall mechanical properties relative to free-swelling controls. Equilibrium modulus values reach 1306+/-79 kPa and glycosaminoglycan levels reach 8.7+/-1.6% w.w. during this 8-week period and are similar to host cartilage properties (994+/-280 kPa, 6.3+/-0.9% w.w.). CONCLUSIONS: An optimal strategy for the functional tissue engineering of articular cartilage, particularly to accelerate construct development, may incorporate sequential application of different growth factors and applied deformational loading.

Glucosamine promotes chondrogenic phenotype in both chondrocytes and mesenchymal stem cells and inhibits MMP-13 expression and matrix degradation.

OBJECTIVES: Glucosamine (GlcN), a natural amino monosaccharide, is a constituent of glycosaminoglycans (GAGs) found in hyaline cartilage. GlcN salts constitute a new class of nutraceutical components with putative chondroprotective activity, which may target chondrocytes as well as chondroprogenitors cells, such as mesenchymal stem cells (MSCs), during cartilage turnover and repair. In the present study, we examined the effects of GlcN on chondrogenesis of human MSCs (hMSCs) and the phenotype of normal and osteoarthritic human articular chondrocytes, using an in vitro pellet culture model maintained in a defined medium. METHODS: hMSCs and normal and osteoarthritic human chondrocytes grown as pellet cultures, stimulated or not with interleukin-1beta (IL-1beta), were treated with varying doses of GlcN. Expression of cartilage matrix genes and cartilage degrading enzymes was determined by semiquantitative and quantitative real-time reverse transcription polymerase chain reaction (RT-PCR), and by histological staining of cartilage markers, as well as sulfated GAG (sGAG) analysis and Western blotting. RESULTS: Chondrocytes grown in the presence of serum for 11 days showed decreased expression of the cartilage matrix genes, collagen type II (collagen II) and aggrecan, as early as day 3, which was reversed with GlcN treatment by day 11. Both hMSCs and chondrocytes grown as pellet cultures in defined medium and treated with 100 microM GlcN exhibited enhanced expression of collagen II and aggrecan as well as increased content of sGAG, when compared to control untreated pellets. However, high doses of GlcN (10-20mM) were inhibitory. GlcN treatment partially blocked IL-1beta mediated downregulation of collagen II and aggrecan expression and inhibited expression of the matrix degrading enzyme, matrix metalloproteinase 13 (MMP-13), in both chondrocytes and hMSCs undergoing chondrogenesis. CONCLUSIONS: These observations suggest that GlcN treatment enhances hMSC chondrogenesis and maintains cartilage matrix gene expression in chondrocytes, which may account for some of the reported chondroprotective properties of GlcN on cartilage.

Regulation of cartilaginous ECM gene transcription by chondrocytes and MSCs in 3D culture in response to dynamic loading.

This study explored the biologic response of chondrocytes and mesenchymal stem cells (MSCs) to a dynamic mechanical loading regime. We developed a time-efficient methodology for monitoring regional changes in extracellular matrix gene transcription using reporter promoter constructs. Specifically, transfected cells were homogenously distributed throughout agarose hydrogel constructs, and spatial and temporal gene expression and the ability to form functional ECM were analyzed in response to dynamic mechanical stimuli. Theoretical analyses were used to predict the physical signals generated within the gel in response to these loading regimes. Using a custom compression bioreactor system, changes in aggrecan and type II collagen promoter activity in transfected chondrocyte-laden cylindrical constructs were evaluated in response to a range of loading frequencies and durations. In general, aggrecan promoter activity increased with increasing duration of loading, particularly in the outer annulus region. Interestingly, type II collagen promoter activity decreased in this annular region under identical loading conditions. In addition, we explored the role of mechanical compression in directing chondrogenic differentiation of MSCs by monitoring short-term aggrecan promoter activity. As an example of long-term utility, a specific loading protocol was applied to MSC-laden constructs for 5 days, and the resultant changes in glycosaminoglycan (GAG) production were evaluated over a 4-week period. This dynamic loading regime increased not only short-term aggrecan transcriptional activity but also GAG deposition in long-term culture. These results demonstrate the utility of a new reporter promoter system for optimizing loading protocols to improve the outcome of engineered chondrocyte- and MSC-laden cartilaginous constructs.

Chondrogenic differentiation and functional maturation of bovine mesenchymal stem cells in long-term agarose culture.

BACKGROUND: The developmental history of the chondrocyte results in a cell whose biosynthetic activities are optimized to maintain the concentration and organization of a mechanically functional cartilaginous extracellular matrix. While useful for cartilage tissue engineering studies, the limited supply of healthy autologous chondrocytes may preclude their clinical use. Consequently, multipotential mesenchymal stem cells (MSCs) have been proposed as an alternative cell source. OBJECTIVE: While MSCs undergo chondrogenesis, few studies have assessed the mechanical integrity of their forming matrix. Furthermore, efficiency of matrix formation must be determined in comparison to healthy chondrocytes from the same donor. Given the scarcity of healthy human tissue, this study determined the feasibility of isolating bovine chondrocytes and MSCs, and examined their long-term maturation in three-dimensional agarose culture. EXPERIMENTAL DESIGN: Bovine MSCs were seeded in agarose and induced to undergo chondrogenesis. Mechanical and biochemical properties of MSC-laden constructs were monitored over a 10-week period and compared to those of chondrocytes derived from the same group of animals maintained similarly. RESULTS: Our results show that while chondrogenesis does occur in MSC-laden hydrogels, the amount of the forming matrix and measures of its mechanical properties are lower than that produced by chondrocytes under the same conditions. Furthermore, some important properties, particularly glycosaminoglycan content and equilibrium modulus, plateau with time in MSC-laden constructs, suggesting that diminished capacity is not the result of delayed differentiation. CONCLUSIONS: These findings suggest that while MSCs do generate constructs with substantial cartilaginous properties, further optimization must be done to achieve levels similar to those produced by chondrocytes.

A three-dimensional nanofibrous scaffold for cartilage tissue engineering using human mesenchymal stem cells.

The utilization of adult stem cells in tissue engineering is a promising solution to the problem of tissue or organ shortage. Adult bone marrow derived mesenchymal stem cells (MSCs) are undifferentiated, multipotential cells which are capable of giving rise to chondrocytes when maintained in a three-dimensional culture and treated with members of the transforming growth factor-beta (TGF-beta) family of growth factors. In this study, we fabricated a nanofibrous scaffold (NFS) made of a synthetic biodegradable polymer, poly(-caprolactone) (PCL), and examined its ability to support in vitro chondrogenesis of MSCs. The electrospun PCL porous scaffold was constructed of uniform, randomly oriented nanofibers with a diameter of 700 nm, and structural integrity of this scaffold was maintained over a 21-day culture period. MSCs cultured in NFSs in the presence of TGF-beta1 differentiated to a chondrocytic phenotype, as evidenced by chondrocyte-specific gene expression and synthesis of cartilage-associated extracellular matrix (ECM) proteins. The level of chondrogenesis observed in MSCs seeded within NFSs was comparable to that observed for MSCs maintained as cell aggregates or pellets, a widely used culture protocol for studying chondrogenesis of MSCs in vitro. Due to the physical nature and improved mechanical properties of NFSs, particularly in comparison to cell pellets, the findings reported here suggest that the PCL NFS is a practical carrier for MSC transplantation, and represents a candidate scaffold for cell-based tissue engineering approaches to cartilage repair.

Adult mesenchymal stem cells: characterization, differentiation, and application in cell and gene therapy.

A considerable amount of retrospective data is available that describes putative mesenchymal stem cells (MSCs). However, there is still very little knowledge available that documents the properties of a MSC in its native environment. Although the precise identity of MSCs remains a challenge, further understanding of their biological properties will be greatly advanced by analyzing the mechanisms that govern their self-renewal and differentiation potential. This review begins with the current state of knowledge on the biology of MSCs, specifically with respect to their existence in the adult organism and postulation of their biological niche. While MSCs are considered suitable candidates for cell-based strategies owing to their intrinsic capacity to self-renew and differentiate, there is currently little information available regarding the molecular mechanisms that govern their stem cell potential. We propose here a model for the regulation of MSC differentiation, and recent findings regarding the regulation of MSC differentiation are discussed. Current research efforts focused on elucidating the mechanisms regulating MSC differentiation should facilitate the design of optimal in vitro culture conditions to enhance their clinical utility cell and gene therapy.

Mesenchymal stem cell-based cartilage tissue engineering: cells, scaffold and biology.

Cartilage repair and regeneration by stem cell-based tissue engineering could be of enormous therapeutic and economic potential benefit for an aging population. However, to use stem cells effectively, their natural environment must be understood in order to expand them in vitro without compromising their multilineage potential and their specific differentiation program. Collaboration between diverse academic disciplines and between research and regulatory government agencies and industry is crucial before cell-based cartilage tissue engineering can achieve its full therapeutic potential.

Estrogenic endocrine disruptive components interfere with calcium handling and differentiation of human trophoblast cells.

During development, calcium (Ca) is actively transported by placental trophoblasts to meet fetal nutritional and the skeletal mineralization needs. Maternal exposure to estrogenic pesticides, such as 1,1-bis(p-chlorophenyl)-2,2,2-trichloroethane (DDT) and methoxychlor (MTC), has been shown to result in reproductive disorders and/or abnormal fetal development. In this study, we have examined the effects of exposure of trophoblastic cells to MTC and DTT, in comparison to 17beta-estradiol (E2) and diethylstilbestrol (DES), to test the hypothesis that cellular Ca handling is a target for these endocrine disruptive components. Treatment with DDT, MTC, DES, or E2 increased cellular Ca uptake, and the expression of trophoblast-specific human Ca binding protein (HCaBP) was down-regulated by both MTC and DDT. Treatment with MTC, DDT, and DES inhibited cell proliferation, induced apoptosis, and suppressed expression of several trophoblast differentiation marker genes. These effects were reversed by overexpression of metallothionein IIa, a gene highly responsive to cadmium and other metals. These results strongly suggest that trophoblast Ca handling functions are endocrinally modulated, and that their alteration by candidate endocrine disruptors, such as MTC and DDT, constitutes a possible pathway of the harmful effects of these components on fetal development.