Actin mRNA Research Articles

Effect of histone deacetylase inhibitors on heat shock protein gene expression during Xenopus development.

We examined the effect of histone deacetylase inhibitors (HDIs), trichostatin A (TSA), valproic acid (VPA), and sodium butyrate (NaB) on heat shock protein (hsp) gene expression during early Xenopus laevis development. HDIs enhance histone acetylation and result in the relief of repressed chromatin domains and ultimately increase the accessibility of transcription factors to target cis-acting regulatory sites. Treatment of embryos with HDIs enhanced the heat shock-induced accumulation of hsp70 mRNA in post-midblastula stage embryos. No effect was observed with actin mRNA or other hsp70 family members including heat shock cognate 70 and immunoglobulin binding protein. Normally, hsp30 genes are not heat-inducible until the late neurula or early tailbud stage of development. Treatment with HDIs resulted in heat-induced expression of hsp30 genes at the gastrula stage with enhanced heat-induced accumulation in neurula and tailbud stages. HDI treatment alone did not induce the accumulation of hsp70 or hsp30 mRNA. Whole-mount in situ hybridization verified the RNA blot analyses and additionally revealed that TSA treatment did not result in any major alterations in the spatial pattern of stress-induced hsp70 or hsp30 mRNA accumulation in early embryos. This study suggests that the states of Xenopus hsp70 and 30 chromatin are subject to repression beyond the midblastula transition.

Apoptotic Body Engulfment by a Human Stellate Cell Line Is Profibrogenic

Hepatocyte apoptosis and stellate cell activation are both features of chronic liver diseases, but a relationship between these events has not been explored. In macrophages, engulfment of apoptotic bodies induces expression of transforming growth factor-β (TGF-β), a profibrogenic cytokine. We examined whether a similar response occurs in stellate cells. Fluorescently labeled hepatocyte apoptotic bodies were added to cultures of primary and immortalized human stellate cells. Stellate cells, but not hepatocytes, readily engulfed apoptotic bodies in a time-dependent manner as assessed by confocal microscopy. The activation of primary and immortalized human stellate cells after incubation with apoptotic bodies, as well as their fibrogenic activity, was indicated by an increase in α-smooth muscle actin (primary cells), TGF-β1, and collagen α1(I) mRNA (primary and immortalized cells). The profibrogenic response was dependent upon apoptotic body engulfment, because nocodazole, a microtubule-inhibiting agent, blocked both the engulfment and the increase of TGF-β1 and collagen α1(I) mRNA. As described in primary rodent stellate cells, up-regulation of collagen α1(I) mRNA was inhibited by a PI-3K inhibitor (LY294002) and a p38 mitogen-activated protein kinase inhibitor (SB203580) in LX-1 cells. In conclusion, these data support a model in which engulfment of hepatocyte apoptotic bodies by stellate cells leads to a fibrogenic response by eliciting a kinase-signaling pathway.

Atrophy responses to muscle inactivity. I. Cellular markers of protein deficits.

The goal of this study was to use the model of spinal cord isolation (SI), which blocks nearly all neuromuscular activity while leaving the motoneuron muscle-fiber connections intact, to characterize the cellular processes linked to marked muscle atrophy. Rats randomly assigned to normal control and SI groups were studied at 0, 2, 4, 8, and 15 days after SI surgery. The slow soleus muscle atrophied by approximately 50%, with the greatest degree of loss occurring during the first 8 days. Throughout the SI duration, muscle protein concentration was maintained at the control level, whereas myofibrillar protein concentration steadily decreased between 4 and 15 days of SI, and this was associated with a 50% decrease in myosin heavy chain (MHC) normalized to total protein. Actin relative to the total protein was maintained at the control level. Marked reductions occurred in total RNA and DNA content and in total MHC and actin mRNA expressed relative to 18S ribosomal RNA. These findings suggest that two key factors contributing to the muscle atrophy in the SI model are 1). a reduction in ribosomal RNA that is consistent with a reduction in protein translational capacity, and 2). insufficient mRNA substrate for translating key sarcomeric proteins comprising the myofibril fraction, such as MHC and actin. In addition, the marked selective depletion of MHC protein in the muscles of SI rats suggests that this protein is more vulnerable to inactivity than actin protein. This selective MHC loss could be a major contributor for the previously reported loss in the functional integrity of SI muscles. Collectively, these data are consistent with the involvement of pretranslational and translational processes in muscle atrophy due to SI.

Quantification of multiple mRNA levels in rat brain regions using real time optical PCR

TaqMan ® is significantly more sensitive than other methods of mRNA quantification and makes possible the simultaneous analysis of numerous genes in small brain regions. This technique was used to quantify levels of mRNAs of 21 genes in tissue extracts from caudate putamen and nucleus accumbens from individual rats after 1 day ‘binge’ cocaine or saline administration. Expression of glyceraldehyde-3-phosphate-dehydrogenase, cyclophilin and actin mRNAs as well as 18S ribosomal RNA were evaluated for normalization of levels of gene expression.

Contractile activity of human decidual stromal cells.

We previously demonstrated that human decidual stromal cells (DSC), the main cellular component of the decidua, are similar in antigen phenotype and structure to myofibroblasts, cells with contractile activity. In this work we isolated and maintained DSC in fibroblast medium, in which these cells show a stable phenotype similar to that of DSC in vivo. Flow cytometric observations showed that most DSC expressed alpha-smooth muscle (alpha-SM) actin, an intermediate filament that is considered a marker of myofibroblasts and is responsible for the contractile activity of these cells. alpha-SM actin mRNA was detected by RT-PCR in these cells. The contractile activity of DSC was determined by the gel contraction assay; we found that TGF beta 1 and platelet-derived-growth factor, cytokines that are known to be inducers of myofibroblast contractility, also induced contractility of DSC. IL-2, a Th1 cytokine-related with spontaneous abortion, also activated DSC contractility. Our results confirmed that DSC are phenotypically and functionally related with myofibroblast.

Trypanosoma cruzi: allelic comparisons of the actin genes and analysis of their transcripts

Two allelic genomic fragments containing actin genes from Trypanosoma cruzi were isolated and characterized. One allele comprises two complete tandem repeats of a sequence encoding an actin gene. In the other, only one actin gene is found. Each one of these three gene copies encode for a complete and identical potential protein of 376 amino acids which is 93% similar with its homolog from Trypanosoma brucei. Northern hybridizations of both total and polysomal RNA from epimastigotes demonstrated the presence of an actin polyadenylated mRNA of about 1.6 kb. Actin transcripts processing sites were determined by 5 ′- and 3 ′-RACE. The obtained sequence data demonstrates that actin genes from both alleles are expressed. The stability of actin mRNA was found to be similar to the one exhibited by the ribosomal protein S4 mRNA as an internal reference. A time course analysis of cultured epimastigotes showed a novel behaviour in which actin mRNA steady state concentration peaks during the transition from the logarithmic to the stationary phase of growth. Index Descriptors and Abbreviations: cDNA, complementary DNA; EDTA, ethylenediaminetetraacetic acid; mRNA, messenger RNA; ORF, open reading frame; PCR, polymerase chain reaction; RACE, rapid amplification of cDNA ends; RNase, ribonuclease; SIRE, short interspersed element; ssrRNA, small subunit ribosomal RNA; UV, ultraviolet; UTR, untranslated region

Diagnosis of dermatophytoses: conventional methods and recent molecular biological methods

Dermatophytoses such as tinea pedis and tinea unguium are very common diseases in the field of dermatology. The diagnosis of dermatophytoses is usually performed by direct microscopy and culture. The identification of species is based on morphological features of giant culture and slide culture. However, in some cases, it is difficult to identify the species clearly because the culture shows an atypical appearance or is false negative. Therefore, several molecular biological methods have been developed for precise identification of a species. The analysis of patterns of random amplification of polymorphic DNA (RAPD) and restriction fragment length polymorphisms (RFLP) of mitochondrial DNA is useful for identifying isolates which are not clearly identifiable by conventional biological techniques. The phylogenetic analysis of dermatophytes was made by using DNA direct sequencing of nuclear ribosomal internal transcribed spacer 1 (ITS1). Sequence analysis of chitin synthase 1 (CHS 1) is a rapid tool for species level identification. We attempted the identification and viability assessment of dermatophytes based on the quantitative measurement of dermatophyte actin (ACT) mRNA. An internal fragment of the ACT, 725 to 762 bp, was isolated by PCR from the genomic DNA of dermatophytes and sequenced. ACT intron based primers were dermatophyte species-specific and primer pairs crossing the intron were dermatophyte genus-specific. The results indicated that quantification of dermatophyte ACT mRNA correlated with the results of culture and KOH examination. It is important that the identification of dermatophyte be done by combining conventional methods with molecular biological methods. In some cases results of the two methods do not correspond, and is those the fungal species needs to be re-examined.

Effect of Endocrine Disrupters on mRNA Expression of Vitellogenin (VTG) II and Very Low Density Lipoprotein (apoVLDL) II in the Liver of Quail Embryos.

The present study was conducted to assess estrogenic activity of nonylphenol (NP) and octylphenol (OP) in quail embryos by determining mRNA levels of liver vitellogenin (VTG) II and very low density lipoprotein (apoVLDL) II. The fertile eggs were treated with a single injection of either NP, OP or ethynyl estradiol (EE) at doses of 10 and 100 nmole/egg in 20μl on day 13 of incubation. In the control group the eggs were treated with the vehicle (corn oil, 20μl/egg). On day 15 of incubation the liver was collected and total RNA was extracted. Both mRNA levels were determined by RT-PCR assay and were expressed in relation to β actin mRNA levels. No expression of VTG II mRNA was detected in the control group, whereas a marked induction of VTG II mRNA was revealed in the EE treatment. A weak but distinct expression of VTG II mRNA was evident in the NP and OP treatment groups. ApoVLDLII transcripts were detected in the control group and induced markedly by the injection of EE with higher expression in females. NP also induced considerable expression in females, whereas no transcripts were detected in males. OP also induced the transcript in females but in males OP at 10 nmole was effective. This study indicates that NP and OP possess estrogenic activity in terms of liver VTG II and apoVLDLII mRNA expression in the quail embryo, and that apoVLDLII expression in female embryo is more sensitive to estrogenic substances.

RNA travels with ZBP1

On page 77, Farina et al. get a handle on RNA localization machinery by proving that an RNA-binding protein is essential for both mRNA transport and cell motility. Figure Lamella-localized ZBP1 (white) granules do not form if ZBP1 cannot bind actin mRNA (right). The cell motility connection comes about because localization of the β-actin mRNA to the lamellae is required for cell polarity and motility in fibroblasts. The new results show that this localization depends on ZBP1, a protein associated with cytoplasmic granules that contain the actin mRNA. ZBP1 bound to the mRNA through two COOH-terminal KH domains that were required for granule formation and attachment to the actin cytoskeleton. NH2-terminal regions of ZBP1 were necessary for granule localization in the lamellae. Dominant–negative ZBP1 constructs that mislocalized actin RNA inhibited fibroblast motility. Since mRNAs for some actin-associated proteins, such as ARP3, also contain ZBP1-binding sequences, the authors believe that ZBP1 may link several messages involved in motility to a transport complex. But ZBP1 may be more than just a scaffold: a ZBP1 homologue has been linked to translational repression of the insulin-related growth factor. Perhaps ZBP1 also ensures that the actin mRNA is not translated until it reaches its ultimate destination.The group plans to purify ZBP1- associated proteins in the complex to identify the motor responsible for actin-based transport. ZBP1 is also known to be associated with microtubules in neurons, so it may connect to different motors depending on the cell type. ▪

Identification of differentially expressed genes like cofilin2 in growing collateral arteries

Arteriogenesis, the growth of pre-existing collateral arteries, can be induced in rabbits by occlusion of the femoral artery. In order to analyze the differential gene expression in arteriogenesis, cDNA of collateral arteries 24 h after femoral occlusion or sham operation was subjected to suppression subtractive hybridization (SSH). We demonstrated an upregulation of the U6 snRNA binding protein Lsm5, cytochrome b, an expressed sequence tag, and the actin-depolymerizing factor cofilin2 mRNA in collateral arteries 24 h after femoral ligation. For cofilin2, we also detected an increase in the protein level and a localization predominantly in smooth muscle cells of collaterals. Simultaneously with the upregulation of cofilin2 we found a downregulation of the α-smooth muscle actin mRNA in growing collateral arteries. In summary, our data showed an augmented expression level of genes contributing to different fundamental processes of arteriogenesis.

The cytoplasmic polyadenylation element binding protein and polyadenylation of messenger RNA in Aplysia neurons

Translation of some mRNAs in nerve terminals has been shown to be regulated by polyadenylation in an experience-dependent manner. The transcripts whose translation is controlled by regulated polyadenylation contain the cytoplasmic polyadenylation element (CPE), which binds to the highly conserved CPE-binding protein (CPEB). In Aplysia, neuron-specific actin mRNA, which has a CPE in its 3′ UTR, is located both in cell bodies and at nerve endings (synaptosomes). We found that actin mRNA from pleural ganglion sensory neurons becomes polyadenylated during long-term facilitation produced by treatment with serotonin or 8-bromo cAMP. We cloned two isoforms of CPEB (ApCPEB77 and ApCEPB49) from Aplysia nervous tissue. The larger form, which is predominant in nervous tissue, is similar to p82, the clam binding protein, as well as to vertebrate CPEBs. Moreover, synaptosomal actin mRNAs are polyadenylated following the treatment with 5-HT. Since both CPEB and polyadenylated actin mRNA are present in synaptosomes and synaptosomal actin protein increases during long-term facilitation, we suggest that the translation of actin message in nerve endings is up-regulated by polyadenylation to grow new synapses.

Enhanced accumulation of constitutive heat shock protein mRNA is an initial response of eye tissue to mild hyperthermia in vivo in adult Xenopus laevis.

We have examined the effect of mild hyperthermia in vivo on heat shock transcription factor (HSF) binding activity and heat shock protein (hsp) gene expression in eye tissue of adult Xenopus laevis. A specific interaction between HSF and a synthetic oligonucleotide corresponding to the proximal heat shock element of the Xenopus hsp70B gene was greatly enhanced in eyes from hyperthermic animals compared with controls. Given these results, we examined the effect of hyperthermia in vivo on the expression of five hsp genes (hsp70, hsc70, BiP, hsp90, and hsp30) in eye tissue. Interestingly, at 28 degrees C constitutively expressed hsp genes hsc70, BiP, and hsp90 were strongly enhanced, with further accumulation at 30 degrees C. However, hsp70 and hsp30 mRNA accumulation were not detectable at 28 degrees C but were strongly induced at 30 degrees C. No enhancement of the relative levels of cytoskeletal actin mRNA was observed in the eye tissue of hyperthermic animals. These results suggest that one of the primary responses of eye tissue to hyperthermia in vivo is in the elevation of mRNAs encoding a set of constitutively expressed molecular chaperones.

The effects of feeding conjugated linoleic acid on pig liver vitamin A and retinol binding protein mRNA

Banni et al. (1999) indicated feeding conjugated linoleic acid (CLA) increases rat liver retinol and retinyl-ester levels. We wanted to determine if feeding CLA would affect pig vitamin A status. Feeding 0.5% CLA did not increase pig liver retinyl-palmitate but did increase retinol from 1.56 to 2.56 109g g-1 (P < 0.05) and the level of retinol binding protien mRNA relative to actin mRNA (P < 0.05). Key words: Conjugated linoleic acid, pig, vitamin A, retinol, retinol binding protein

Mice lacking skeletal muscle actin show reduced muscle strength and growth deficits and die during the neonatal period.

All four of the muscle actins (skeletal, cardiac, vascular, and enteric) in higher vertebrates show distinct expression patterns and display highly conserved amino acid sequences. While it is hypothesized that each of the muscle isoactins is specifically adapted to its respective tissue and that the minor variations among them have developmental and/or physiological relevance, the exact functional and developmental significance of these proteins remains largely unknown. In order to begin to assess these issues, we disrupted the skeletal actin gene by homologous recombination. All mice lacking skeletal actin die in the early neonatal period (day 1 to 9). These null animals appear normal at birth and can breathe, walk, and suckle, but within 4 days, they show a markedly lower body weight than normal littermates and many develop scoliosis. Null mice show a loss of glycogen and reduced brown fat that is consistent with malnutrition leading to death. Newborn skeletal muscles from null mice are similar to those of wild-type mice in size, fiber type, and ultrastructural organization. At birth, both hemizygous and homozygous null animals show an increase in cardiac and vascular actin mRNA in skeletal muscle, with no skeletal actin mRNA present in null mice. Adult hemizygous animals show an increased level of skeletal actin mRNA in hind limb muscle but no overt phenotype. Extensor digitorum longus (EDL) muscle isolated from skeletal-actin-deficient mice at day 2 to 3 showed a marked reduction in force production compared to that of control littermates, and EDL muscle from hemizygous animals displayed an intermediate force generation. Thus, while increases in cardiac and vascular smooth-muscle actin can partially compensate for the lack of skeletal actin in null mice, this is not sufficient to support adequate skeletal muscle growth and/or function.

Telomerase immortalization of human myometrial cells.

Several strategies have been described for the primary culture of human myometrial cells. However, primary cultures of myometrial cells have a limited life span, making continual tissue acquisition and cell isolation necessary. Recent studies have demonstrated that cell culture life span is related to chromosomal telomere length, and cellular senescence results from progressive telomere shortening and the lack of telomerase expression. Transfection of cells with expression vectors containing the human telomerase reverse transcriptase (hTERT) maintains telomere length and effectively gives normal cells an unlimited life span in culture. In addition, hTERT extends the life span of cultured cells far beyond normal senescence without causing neoplastic transformation. In the present study, we developed a cell line from hTERT-infected myometrial cells (hTERT-HM). Cells were isolated from myometrial tissue obtained from women undergoing hysterectomy, and retroviral infection was used to express the catalytic subunit of telomerase in myometrial cells. Cells expressing hTERT have been in continuous culture for >10 mo, whereas the control culture senesced after approximately 2 mo. Telomerase activity was monitored in cells with a polymerase chain reaction-based telomerase activity assay. Telomerase-expressing cells contained mRNA for alpha smooth muscle actin, smoothelin, oxytocin receptor, and estrogen receptor alpha, but the estrogen receptor beta receptor was lost. Immunoblotting analysis identified the expression of calponin, caldesmon, alpha smooth muscle actin, and oxytocin receptor. Although estrogen receptor expression was below the level of detection with immunoblotting, transfection experiments performed with reporter constructs driven by estrogen response elements demonstrated estrogen responsiveness in the hTERT-HM. In addition, treatment of hTERT-HM with oxytocin caused a concentration-dependent increase in intracellular calcium levels, confirming the presence of functional oxytocin receptors. Myometrial cells immortalized with hTERT retained markers of differentiation that are observed in primary cultures of smooth muscle cells. The expression of various smooth muscle/myometrium cell markers suggests that these cells may be an appropriate model system to study certain aspects of human myometrial function.

NF-kappaB induced by IL-1beta inhibits elastin transcription and myofibroblast phenotype.

Interleukin (IL)-1beta released after lung injury regulates the production of extracellular matrix components. We found that IL-1beta treatment reduced the rate of elastin gene transcription by 74% in neonatal rat lung fibroblasts. Deletion analysis of the rat elastin promoter detected a cis-acting element located at -118 to -102 bp that strongly bound Sp1 and Sp3 but not nuclear factor (NF)-kappaB. This element mediated IL-1beta-induced inhibition of the elastin promoter. IL-1beta treatment did not affect the level of Sp1 but did induce translocation of the p65 subunit of NF-kappaB. Overexpression of p65 decreased elastin promoter activity and markedly reduced elastin mRNA. Immunoprecipitation studies indicated an interaction between the p65 subunit and Sp1 protein. Microarray analysis of mRNA isolated after overexpression of p65 or treatment with IL-1beta revealed downregulation of alpha-smooth muscle actin and calponin mRNAs. Expression of these genes is associated with the myofibroblast phenotype. These results indicate that IL-1beta activates the nuclear localization of NF-kappaB that subsequently interacts with Sp1 to downregulate elastin transcription and expression of the myofibroblast phenotype.

Expression of fetal isoforms of actin after transplantation injury

Trauma and injury to transplanted organs in the early post-transplant period are significant factors that affect long-term graft survival. Fetal isoforms of actin are integral members of the immediate early gene family and are expressed in response to free radical injury. We therefore studied actin gene expression in heart transplantation to determine if reperfusion injury activates fetal isoforms of actins. Heterotopic cardiac transplantations were performed in mice. mRNA was extracted from allo- and isografted hearts as well as from normal hearts and spleen. Northern hybridization with actin cDNA to alpha and beta/gamma actin mRNA was performed and analyzed by densitometry. The beta/gamma actin gene expression in the transplanted hearts was found to be significantly elevated within 48 h after transplantation. Analysis of beta/gamma actin gene expression in isografts substantiates the possibility of de novo increase in actin expression. Our studies demonstrate for the first time that fetal isoforms of actin are induced in the allograft heart after transplantation.

Expression of fetal isoforms of actin after transplantation injury.

Trauma and injury to transplanted organs in the early post-transplant period are significant factors that affect long-term graft survival. Fetal isoforms of actin are integral members of the immediate early gene family and are expressed in response to free radical injury. We therefore studied actin gene expression in heart transplantation to determine if reperfusion injury activates fetal isoforms of actins. Heterotopic cardiac transplantations were performed in mice. mRNA was extracted from allo- and isografted hearts as well as from normal hearts and spleen. Northern hybridization with actin cDNA to alpha and beta/gamma actin mRNA was performed and analyzed by densitometry. The beta/gamma actin gene expression in the transplanted hearts was found to be significantly elevated within 48 h after transplantation. Analysis of beta/gamma actin gene expression in isografts substantiates the possibility of de novo increase in actin expression. Our studies demonstrate for the first time that fetal isoforms of actin are induced in the allograft heart after transplantation.

Metastasis-inducing dna regulates the expression of the osteopontin gene by binding the transcription factor Tcf-4.

Small 1,000-bp fragments of genomic DNA obtained from human malignant breast cancer cell lines when transfected into a benign rat mammary cell line enhance transcription of the osteopontin gene and thereby cause the cells to metastasize in syngeneic rats. To identify the molecular events underlying this process, transient cotransfections of an osteopontin promoter-reporter construct and fragments of one metastasis-inducing DNA (Met-DNA) have identified the active components in the Met-DNA as the binding sites for the T-cell factor (Tcf) family of transcription factors. Incubation of cell extracts with active DNA fragments containing the sequence CAAAG caused retardation of their mobilities on polyacrylamide gels, and Western blotting identified Tcf-4, beta-catenin, and E-cadherin in the relevant DNA complexes in vitro. Transfection of an expression vector for Tcf-4 inhibited the stimulated activity of the osteopontin promoter-reporter construct caused by transiently transfected active fragments of Met-DNA or permanently transfected Met-DNA. This stimulated activity of the osteopontin promoter-reporter construct is accompanied by an increase in endogenous osteopontin mRNA but not in fos or actin mRNAs in the transfected cells. Permanent transfection of the benign rat mammary cell line with a 20-bp fragment from the Met-DNA containing the Tcf recognition sequence CAAAG caused an enhanced permanent production of endogenous osteopontin protein in vitro and induced the cells to metastasize in syngeneic rats in vivo. The corresponding fragment without the CAAAG sequence was without either effect. Therefore, the regulatory effect of the C9-Met-DNA is exerted, at least in part, by a CAAAG sequence that can sequester the endogenous inhibitory Tcf-4 and thereby promote transcription of osteopontin, the direct effector of metastasis in this system.

Monoamine Oxidase Expression and Activity in Human Placentae from Pre-eclamptic and Normotensive Pregnancies

A feature of pre-eclampsia is that circulating levels of maternal serotonin (5-hydroxytryptamine) are elevated and placental monoamine oxidase-A (MAO-A) activity, the major factor in the regulation of serotonin levels in pregnancy, is reduced. It is not known whether this is due to a reduced MAO-A protein content or a reduced catalytic turnover of the serotonin by MAO-A; this question has been addressed in the present work. Term placentae from normotensive and pre-eclamptic women were analysed for MAO-A specific mRNA expression (by semi-quantitative RT-PCR), MAO-A protein (by immunohistochemistry and quantitative ELISA, using a MAO-A specific monoclonal antibody), together with MAO activity (using [3H] labelled 5-hydroxytryptamine as substrate). Immunohistochemical analysis of placentae from both normotensive and pre-eclamptic women demonstrated that MAO-A protein is located in the cytoplasm of the placental syncytiotrophoblast layer, consistent with a mitochondrial location; no MAO-A protein was found in the nucleus. No MAO-B protein was detected in this placental layer, despite the presence of MAO-B mRNA. The results indicate that both total protein/g fresh weight and MAO-A protein/g fresh weight were approximately 40 per cent lower in pre-eclamptic than in normotensive placentae, but that there was no statistical difference in the expression of MAO-A mRNA in relation to GAPDH or actin mRNA or in MAO-A protein/mg total protein. However, MAO-A activity/g fresh weight was significantly reduced in pre-eclamptic placentae, in agreement with previous findings. This was found to be due to a 60 per cent reduction (P< 0.05) in the catalytic turnover (activity/molecule) of the enzyme. This study has therefore clearly shown that the expression of placental MAO-A specific mRNA and MAO-A protein are not specifically affected in pre-eclampsia, but that the catalytic efficiency of the expressed MAO-A enzyme in pre-eclamptic placentae is greatly reduced.