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Molecular heterogeneity of sporadic adult Burkitt-type leukemia/lymphoma as revealed by PCR and cytogenetics: correlation with morphology, immunology and clinical features.

Chromosomal translocations involving the MYC oncogene are a hallmark of Burkitt lymphoma but they are only found in a varying frequency in mature Burkitt-type acute lymphoblastic leukemia (B-ALL). We have investigated samples of 56 sporadic Burkitt leukemia/lymphoma patients for the translocations t(8;14)(q24;q32), t(2;8)(p11;q24) and t(8;22)(q24;q11). Long PCR was used for detecting the immunoglobulin heavy chain (IgH) translocation and cytogenetics and/or fluorescence in situ hybridization for detecting the 'variant' MYC translocations. A total of 29 samples (51.8%) were t(8;14)-positive by long PCR. Approximately one-third had a chromosomal breakpoint in the IgH joining region while the others had breakpoints in the IgH switch regions. Among them were two cases with a previously unreported MYC translocation into the IgE switch region. Long PCR was more reliable compared to conventional cytogenetics for detecting the t(8;14). Epstein-Barr virus was detected in high copy number in two (3.6%) t(8;14)-positive cases by real-time quantitative PCR. Human herpesvirus 8 was not detected in any case by nested PCR. A typical L3 or L3-compatible cytomorphology was highly predictive (>80%) but not specific of a MYC translocation. A total of 34 patients were treated according to the GMALL B-ALL therapy protocols and there was no significant difference in overall survival between patients with or without t(8;14).

Adolescent↗

Adiponectin gene is expressed in multiple tissues in the chicken: food deprivation influences adiponectin messenger ribonucleic acid expression.

Adiponectin is a cytokine hormone originally found to be secreted exclusively by white adipose tissue. However, recent evidences suggest that adiponectin is also produced in brown adipose tissue and skeletal muscle. The present study investigated the expression of adiponectin mRNA in various tissues in the chicken. We also studied the effect of food deprivation on adiponectin gene expression in adipose tissue, liver, anterior pituitary gland, and diencephalon in the chicken. The open reading frame of chicken adiponectin cDNA consists of 735 nucleotides that were 65-68% homologous to various mammalian adiponectin cDNAs. The deduced amino acid sequence of chicken adiponectin contains 22 glycine-X-Y repeats (in which X and Y represent any amino acid) at the N-terminal end as found in the mammalian adiponectin. By RT-PCR and Northern analysis, we detected chicken adiponectin mRNA transcript in adipose tissue, liver, anterior pituitary gland, diencephalon, skeletal muscle, liver, kidney, ovary, and spleen but not in blood. Adiponectin mRNA expression in various tissues was quantitated using real-time quantitative PCR and found to be the highest in adipose tissue, followed by liver, anterior pituitary, diencephalon, kidney, and skeletal muscle. We also found that adiponectin mRNA quantity was significantly decreased after a 48-h food deprivation in adipose tissue, liver, and anterior pituitary gland but not in diencephalon. Our results provide novel evidence that, unlike mammals, adiponectin gene is expressed in several tissues in the chicken and that its expression is influenced by food deprivation.

Adiponectin↗

PCR-based methods for detecting DNA damage and its repair at the sub-gene and single nucleotide levels in cells.

Three PCR-based methods are described that allow covalent drug-DNA adducts, and their repair, to be studied at various levels of resolution from gene regions to the individual nucleotide level in single copy genes. A quantitative PCR (QPCR) method measures the total damage on both DNA strands in a gene region, usually between 300 and 3,000 base pairs in length. Strand-specific QPCR incorporates adaptations that allow damage to be measured in the same region as QPCR but in a strand-specific manner. Single-strand ligation PCR allows the detection of adduct formation at the level of single nucleotides, on individual strands, in a single copy gene in mammalian cells. If antibodies to the DNA adducts of interest are available, these can be used to capture and isolate adducted DNA for use in single-strand ligation PCR increasing the sensitivity of the assay.

Base Sequence↗

Epidemiologic tools for malaria surveillance in an urban setting of low endemicity along the Colombian Pacific coast.

An evaluation of 3 different methods for malaria diagnosis was carried out in an urban area of low endemicity on the Pacific coast of Colombia. Samples were collected from 833 symptomatic patients at a malaria clinic and examined by the polymerase chain reaction (PCR), quantitative buffy coat (QBC; Becton Dickinson, Franklin Lakes, NJ) method, and the traditional thick blood smear. The prevalence of Plasmodium falciparum malaria was 5.88% by thick blood smear, 7.34% by the QBC method, and 21.87% by PCR. The agreement between microscopists was 99.5%. The agreement between the QBC method and thick blood smear was 96.13% (n = 745). Samples positive by PCR but negative by thick blood smear or conversely negative by PCR and positive by thick blood smear were usually of low-level parasitemias. All 3 methods showed agreement in 76.3% of the samples. Sixty-nine (18.8%) samples were positive by PCR but negative by the other 2 methods. Ten samples were positive by both the QBC method and thick blood smear but negative by PCR; most of them had low-level parasitemias. The use of malaria diagnostic methods for epidemiologic surveillance is discussed.

Animals↗

Endothelin in a murine model of cerebral malaria.

Cerebral malaria (CM) remains a deadly complication of Plasmodium falciparum infection, and children are at high risk of developing encephalopathy as a result of CM. This is probably a consequence of the activation of many of the inflammatory cytokines as well as the glial cells and the vascular endothelium in the brain. We have previously demonstrated that there is a striking reduction in cerebral blood flow by magnetic resonance imaging when mice are infected with Plasmodium berghei ANKA (PbA), and we now demonstrate a possible role for endothelin (ET-1) in the pathogenesis of CM. The brains of female C57BL/6 mice with PbA infection were examined at Day 5 for the expression of ET-1, endothelin converting enzyme (ECE), and the endothelin receptors A and B (ET(A) and ET(B)) by both reverse transcription-polymerase chain reaction (RT-PCR) and quantitative real-time PCR. ET-1 and ECE mRNA expression was markedly increased by RT-PCR in PbA-infected mice. Real-time quantitative PCR demonstrated a 3-fold increase in ET-1 (P < 0.05) and a significant increase in ET(A) and ET(B) expression (P < 0.05) in PbA-infected mice. Histopathology bof PbA-infected mice demonstrated a transformation in the morphology of microglial cells and clustering of these cells consistent with activation. Though the full impact of ET-1 on CM remains to be elucidated, these findings demonstrate that in the murine model, there is a significant increase in ET-1 and its components, which is associated with the vasculopathy and immunopathology of CM.

Animals↗

Minimal residual disease after bone marrow transplantation for chronic myelogenous leukemia and implications for graft-versus-leukemia effect: a review of recent results.

Cure of leukemia by allogeneic BMT is achieved by the combined effect of the myeloablative preparative regimen and an allo-immune response of donor cells to residual leukemia termed the graft-versus-leukemia (GVL) effect. In the first year following BMT for CML, PCR used to detect the leukemia-specific BCR/ABL message frequently reveals subclinical levels of persisting leukemia. In a meta-analysis of reports on qualitative PCR findings after BMT for CML in 12 recently published series, we found that for unmanipulated BMT in chronic phase, PCR detection was not associated with a higher relapse risk and that most patients became PCR negative within 2 years post-BMT. In contrast, PCR detection of BCR/ABL transcripts was a more reliable predictor in recipients of T cell-depleted BMT and in those transplanted in accelerated or blastic phase of their disease. For accurate prediction of relapse, serial quantitative PCR is necessary. It could also be used to monitor efficacy of experimental treatments of relapse with interferon or donor lymphocyte transfusions. Furthermore, studies of the association of GVHD with PCR detection of BCR/ABL message may shed light on the relationship of GVL with minimal residual disease in CML.

Bone Marrow Transplantation↗

Detection and quantification of methyl tert-butyl ether-degrading strain PM1 by real-time TaqMan PCR.

The fuel oxygenate methyl tert-butyl ether (MTBE), a widely distributed groundwater contaminant, shows potential for treatment by in situ bioremediation. The bacterial strain PM1 rapidly mineralizes and grows on MTBE in laboratory cultures and can degrade the contaminant when inoculated into groundwater or soil microcosms. We applied the TaqMan quantitative PCR method to detect and quantify strain PM1 in laboratory and field samples. Specific primers and probes were designed for the 16S ribosomal DNA region, and specificity of the primers was confirmed with DNA from 15 related bacterial strains. A linear relationship was measured between the threshold fluorescence (C(T)) value and the quantity of PM1 DNA or PM1 cell density. The detection limit for PM1 TaqMan assay was 2 PM1 cells/ml in pure culture or 180 PM1 cells/ml in a mixture of PM1 with Escherichia coli cells. We could measure PM1 densities in solution culture, groundwater, and sediment samples spiked with PM1 as well as in groundwater collected from an MTBE bioaugmentation field study. In a microcosm biodegradation study, increases in the population density of PM1 corresponded to the rate of removal of MTBE.

Bacteria↗

ADAM28 is overexpressed in human breast carcinomas: implications for carcinoma cell proliferation through cleavage of insulin-like growth factor binding protein-3.

A disintegrin and metalloproteinases (ADAMs) are involved in various biological events including cell adhesion, cell fusion, membrane protein shedding, and proteolysis. In the present study, our reverse transcription-PCR analysis showed that among the 12 different ADAM species with a putative metalloproteinase motif, prototype membrane-anchored ADAM28m and secreted-type ADAM28s are selectively expressed in human breast carcinoma tissues. By real-time quantitative PCR, their expression levels were significantly higher in carcinomas than in nonneoplastic breast tissues. In situ hybridization, immunohistochemistry, and immunoblotting analyses indicated that ADAM28 is predominantly expressed in an active form by carcinoma cells within carcinoma tissues. A direct correlation was observed between mRNA expression levels and proliferative activity of the carcinoma cells. Treatment of ADAM28-expressing breast carcinoma cells (MDA-MB231) with insulin-like growth factor-I (IGF-I) increased cell proliferation, cleavage of IGF binding protein (IGFBP)-3, as well as IGF-I cell signaling; these processes were all significantly inhibited by treatment with ADAM inhibitor or anti-ADAM28 antibody. Down-regulation of ADAM28 expression in MDA-MB231 cells with small interfering RNA significantly reduced cell proliferation, IGFBP-3 cleavage, and growth of xenografts in mice. In addition, cleavage of IGFBP-3 in breast carcinoma tissues was correlated with ADAM28 expression levels and inhibited by treatment with ADAM inhibitor or anti-ADAM28 antibody. These results show that ADAM28 is overexpressed in an activated form in human breast carcinoma cells and suggest that ADAM28 is involved in cell proliferation through enhanced bioavailability of IGF-I released from the IGF-I/IGFBP-3 complex by selective IGFBP-3 cleavage in human breast carcinomas.

ADAM Proteins↗

Strong association of ARK5 with tumor invasion and metastasis.

We recently identified a novel human AMPK family member, ARK5, and discovered that is a major factor in Akt-dependent cancer cell survival and migration activity through activation of MT1-MMPs in vitro. The mRNA expression of other AMPK family members and ARK5 was measured using RT-PCR in human colorectal carcinoma cell lines DLD-1, WiDr, HCT-15, SW620, LoVo, SW480, and mRNA expression of AMPK-alpha1, SNARK, MELK and ARK5, but not AMPK-alpha2, was detected in every line. Quantitative-PCR (Q-PCR) to estimate the amount of ARK5 mRNA expression in the cell lines showed that there is a variety of ARK5 expressions among the cell lines and high expression was observed in a cell line derived from the metastatic lesion, LoVo. To determine the effect of ARK5 overexpression on metastasis in vivo, we established human pancreas cancer cell line PANC-1 stably transfected with ARK5 full-length expression vector (P/ARK) and DLD-1 stably transfected with the same vector (D/ARK). Migration assay showed a remarkable increase in the activity both in P/ARK and D/ARK, and an in vivo metastasis assay showed a marked increase of P/ARK in liver metastasis. Based on these observations, it is suggested that ARK5 expression is involved in cancer invasion and metastasis.

AMP-Activated Protein Kinase Kinases↗

[Carrier and prenatal diagnosis of Duchenne and Becker muscular dystrophy by PCR methods].

The PCR methods for carrier and prenatal diagnosis of Duchenne and Becker muscular dystrophy are described. When a deletion of the dystrophin gene is detected in the proband, the deletion analysis is informative for prenatal diagnosis, and the quantitative PCR analysis of the concerned exons is necessary for carrier diagnosis. When a deletion is not detected, RFLPs analysis should be considered. We analyzed seven polymorphic makers (pERT87-15 combined with digestions with BamHI or XmnI, pERT87-8 combined with digestion with TaqI, and four CA repeat markers at the 5' end and the 3' untranslated regions of the dystrophin gene) by PCR methods. Every woman examined showed a heterozygous finding for at least one of these markers, making it possible to make carrier and prenatal diagnosis. However, it is important to keep in mind that a third of the patients are due to new mutations and that about 10% recombination rate, between the 5' and 3' ends of the dystrophin gene, can exist.

Dystrophin↗

Monitoring of CD95 (APO-1/Fas) ligand expression in human T cells by quantitative RT-PCR.

CD95 (APO-1/Fas) receptor/ligand interaction is a key regulatory pathway for apoptosis in lymphoid cells. We developed a quantitative RT-PCR for the human CD95-L to determine expression levels in lymphoid cell lines and in lymphocytes derived from blood of healthy individuals. In untreated peripheral blood T lymphocytes and T cell lines constitutive expression of the CD95-L mRNA was found at low levels. Stimulation of T cells by treatment with PMA and ionomycine (P/I) lead up to a 100-fold maximal increase in CD95-L mRNA after 4 h. CD95-L mRNA is produced by activated CD8 and CD4T cells. In vivo increased CD95-L mRNA expression was found in freshly isolated T cells during the acute phase of EBV infection. In contrast to T cells, CD95-L mRNA could be induced in some B lineage cell lines only after five days of stimulation. Since defective or accelerated CD95/CD95-L interaction is considered to be involved in the pathogenesis of lymphoproliferation, autoimmunity and AIDS, the quantitative RT-PCR assay described in this paper may provide a powerful tool for monitoring CD95-L expression in these diseases.

Journal Article↗

Frequent infection with a viral pathogen, parvovirus B19, in rheumatic diseases of childhood.

OBJECTIVE: To find further evidence of the association of parvovirus B19 infection with juvenile rheumatic diseases, and to get new insights into the immunopathogenesis of these diseases. METHODS: Paired serum and synovial fluid samples from 74 children with rheumatic disease were analyzed with respect to their content of viral DNA and antibodies directed against the B19 viral proteins VP1, VP2, and NS1. Control sera from 124 children with noninflammatory bone diseases or growth retardation were also analyzed. The sequence of the viral DNA, amplified by polymerase chain reaction (PCR), was determined. IgG-complexed virus was isolated from sera and synovial fluid by adsorption to protein A beads. The amount of free virus versus immunocomplexed virus particles was determined by quantification of the viral genomes by quantitative PCR. RESULTS: Twenty-six of the 74 patients (35%) had detectable amounts of parvovirus B19 DNA in the serum (n = 22 [30%]) and/or the synovial fluid (n = 16 [22%]), whereas only 9 of the 124 control sera (7%) were positive for the viral DNA (P < 0.0001). Forty-six patients (62%) had serum IgG against the structural proteins, indicating past infection with B19. NS1-specific antibodies were detected in sera from 29 patients (39%) and 27 controls (22%) (P < 0.001). In addition, 3 patients (4%) showed VP2-specific IgM. In 15 patients, viral DNA could be repeatedly detected in followup samples of serum and synovial fluid. Sequencing revealed low-degree nucleotide variations that are in the range of genotype 1 of parvovirus B19. Immunocomplexed virus was present in varying amounts, both in the sera and in the synovial fluid samples. CONCLUSION: Parvovirus B19 is frequently found in serum or synovial fluid of children with rheumatism. The rate of persistent B19 infection in these patients is significantly higher than in age-matched controls.

Adolescent↗

Identification of two novel mutations and of a novel critical region in the KRIT1 gene.

Cerebral cavernous malformations (CCMs) represent a common autosomal dominant disorder that predisposes patients to hemorrhagic strokes and focal neurological signs. Mutations in three genes (KRIT1, MGC4607, and PDCD10) have been associated with CCMs. We investigated the role of two new mutations in the KRIT1 gene in two Italian families affected by CCMs. Whole blood DNA was extracted and the mutations were detected after polymerase chain reaction (PCR), denaturing high-performance liquid chromatography screening, and sequencing of the coding regions of the three CCMs-associated genes. Total RNA was extracted, and the KRIT1 cDNA was sequenced and subsequently subjected to real-time quantitative PCR in order to examine the translational outcome of each genomic mutation. A novel splicing acceptor site deletion of the exon 14 in one family and an intronic nucleotide change close to the exon 19 in the other one were identified, both in the KRIT1 gene. These mutations were proven to alter the correct splicing mechanism, resulting, respectively, in a truncated protein of 432 amino acids and in a protein lacking an internal segment. We report two novel cases of splicing affecting genomic variants, suggesting a careful reanalysis of previously identified splice site variations in KRIT1 to look for their possible causative roles of similar missplicing events and their consequent involvement in the pathogenesis of CCMs. Moreover, our genotype-phenotype functional correlation suggests that the C-terminal portion of the KRIT1 protein is likely to contain a short, previously unrecognized segment necessary for its activity.

Adolescent↗

Between prepartum luteolysis and onset of expulsion.

In the cow the foetal endocrine signals that initiate the calving process result in prepartum luteolysis. Withdrawal of progesterone (P4) action is a prerequisite for a normal calving. The rather abrupt declining influence of P4 is followed by a cascade of physiological processes in the myometrium and cervix. This contribution will focus on some of these events. Like in many other species, the myometrium in cows is not completely inactivated during pregnancy. So-called contractures have been registered during the final weeks of gestation and their EMG-characteristics in cows show a low frequency (on average: 13.6 per day) and long duration (on average 12.1 min). They are not evenly spread over the day because they occur less frequently when the cows are disturbed for feeding or cleaning their stables. Contractures affect several foetal functions. In the cow these contractures disappear during a period of about 8-9h when maternal plasma P4 levels are rapidly declining before calving. There is experimental evidence that this temporary inhibition is associated with prepartal luteal regression. The cause of this inhibition is still unknown. Because nitrous oxide inhibits smooth muscle cells and evidence in laboratory animals indicates that expression of the inducible form of nitrous oxide (iNOS) is downregulated in myometrium, but upregulated in the cervix around the onset of parturition, we started to investigate the role of this enzyme in bovine tissues around calving. By means of a RT-PCR technique, we obtained a first indication that iNOS is hardly expressed in the myometrium during calving, while expression was clearly detected at day 4 after calving. Analysis of prepartum en periparturient biopsies from myometrium and cervix with quantitative PCR is still underway. In six pregnant cows, provided with uterine EMG-electrodes and with ultrasonic crystals implanted on the caudal cervical rim to measure cervical dilatation, calving was induced with an injection of prostaglandin (PG) F2alpha. While maternal plasma P4 levels had significantly declined within 8h after PG treatment, the myometrium escaped from temporary inhibition with the development of a parturient contractility pattern on average at 13.5h after injection. However, it was only at 28 h after PG treatment that the first sustained increase of the opening of the vaginal ostium of the cervix was measured.

Animals↗

Predominance of the alpha1D subunit in L-type voltage-gated Ca2+ channels of hair cells in the chicken's cochlea.

The voltage-gated Ca2+ channels that effect tonic release of neurotransmitter from hair cells have unusual pharmacological properties: unlike most presynaptic Ca2+ channels, they are sensitive to dihydropyridines and therefore are L-type. To characterize these Ca2+ channels, we investigated the expression of L-type alpha1 subunits in hair cells of the chicken's cochlea. In PCRs with five different pairs of degenerate primers, we always obtained alpha1D products, but only once an alpha1C product and never an alpha1S product. A full-length alpha1D mRNA sequence was assembled from overlapping PCR products; the predicted amino acid sequence of the alpha1D subunit was about 90% identical to those of the mammalian alpha1D subunits. In situ hybridization confirmed that the alpha1D mRNA is present in hair cells. By using a quantitative PCR assay, we determined that the alpha1D mRNA is 100-500 times more abundant than the alpha1C mRNA. We conclude that most, if not all, voltage-gated Ca2+ channels in hair cells contain an alpha1D subunit. Furthermore, we propose that the alpha1D subunit plays a hitherto undocumented role at tonic synapses.

Amino Acid Sequence↗

Human cell mutants with very low mitochondrial DNA copy number (rho d).

Mutants of human Namalwa cells are described, denoted rho d (mtDNA-depleted), which contain a very low mtDNA copy number. Three isolates, denoted FV02, FV03 and FV05, each selected on the basis of their pyruvate-dependent phenotype, were shown to have severely reduced mitochondrial respiratory functions consistent with results of histochemical and cytochrome analysis. Analyses of mtDNA by the polymerase chain reaction (PCR) and Southern blotting indicated that FV02, FV03 and FV05 each contained a full complement of mtDNA sequences, 16.6 kb in length, in circular form but at a greatly reduced level compared with that of parent rho + cells. Quantitative PCR tests showed first, that the parent Namalwa rho + cells contain about 1800 molecules of mtDNA and second, that cells of each of FV02, FV03 and FV05 contain between 100- and 1000-fold less mtDNA than rho + cells. Subclones of each of FV02, FV03 and FV05 all contained detectable mtDNA, thus eliminating the possibility that these isolates are mixtures of rho o (devoid of mtDNA) and rho + cells. Each of the rho d cell lines FV02, FV03 and FV05 had a more rapid growth rate than the corresponding rho o cells but less than that of the parent Namalwa rho + cells. The experimentally induced rho d cells described here may represent a useful model for human diseases in which severe depletion of cellular mtDNA levels in tissues is encountered.

Cell Division↗

Prospective evaluation of the clinical utility of different methods for the detection of human cytomegalovirus disease after liver transplantation.

Standardized human cytomegalovirus (HCMV) assays were prospectively evaluated to predict HCMV disease. In 135 consecutive adult liver transplantations, pp65-antigenemia, quantitative HCMV-DNA and qualitative pp67-messenger-RNA were determined weekly. No ganciclovir prophylaxis or preemptive treatment was used. One hundred and ten (81.5%) patients showed no HCMV-infection, 25 patients were positive in at least one of the HCMV-tests (18.5%). Four suffered from HCMV viral syndrome (3.0%) and another four from tissue invasive disease. In total, pp65-antigenemia was detected in 18, HCMV-DNA in 22 and pp67-mRNA in 18 patients. The sensitivity and negative predictive value (NPV) for HCMV-disease was 100% for all tests. The PPV for symptomatic HCMV-infection was 47% for pp67 mRNA. In contrast, the PPV of pp65-antigenemia (using a threshold of > 2/200 000 cells) and quantitative PCR (using a cutoff of > 5000 copies/mL) were 80% and 89%, respectively. A cost analysis revealed symptom-triggered or preemptive treatment was less expensive than general ganciclovir prophylaxis, if the incidence of CMV disease was low (<30%). Quantitative human cytomegalovirus (HCMV)-DNA and pp65-antigen assays have a comparable sensitivity and can therefore predict the onset of HCMV symptoms at an early stage. Compared with general prophylaxis, symptom-triggered or preemptive treatment based on one of these assays might reduce the costs and also the danger of ganciclovir resistance.

Antigens, Viral↗

Comparison of human papillomavirus DNA levels in gynecological cancers: implication for cancer development.

We have previously demonstrated the presence of human papillomavirus (HPV) DNA in several gynecological cancers using conventional PCR. In the present study, to further understand the role of HPV in malignant transformation of these cancers, the infection rates and viral loads of HPV 16 and 18 in gynecological cancers were analyzed using real-time quantitative PCR (qPCR). HPV 16 DNA was detected in 61.0% (58/95), 15.2% (7/46) and 32.1% (18/56) of cases of cervical, endometrial and ovarian cancers, respectively. On the other hand, HPV 18 DNA was detected in 23.2% (22/95) of cervical cancers, 1.8% (1/56) of ovarian cancers, and in no cases of endometrial cancer. Thus, HPV 16 is much more prevalent than HPV 18 in malignancies of the female genital tract. We also found that both HPV 16 and 18 were significantly (p < 0.05) less frequently present in endometrial and ovarian cancers than in cervical cancer. The median copy numbers of HPV 16 DNA in endometrial and ovarian cancers were 3,500 and 7,590 copies/microg DNA, respectively. These amounts were also significantly (p < 0.05) lower than HPV 16 DNA in cervical cancer (492,800 copies/microg DNA). Thus, HPV 16 could be detected in all three types of gynecological cancer, whilst HPV 18 is extremely rare in endometrial and ovarian cancers. The lower HPV 16 infection rates and lower copy numbers when compared with cervical cancer tend to suggest that HPV plays a less essential role in the development of endometrial cancer and ovarian cancer.

Adult↗