A pilot study using unique targeted testing of the urogenital microbiome has potential as a predictive test during IVF for implantation outcome.

PURPOSE: This pilot study aimed to develop a methodology characterising the urogenital microbiome as a predictive test in the IVF workup. METHODS: Using unique custom qPCRs, we tested for the presence of specific microbial species from vaginal samples and First Catch Urines from the male. The test panel included a range of potential urogenital pathogens, STIs, 'favourable bacteria' (Lactobacillus spp.) and 'unfavourable bacteria' (anaerobes) reported to influence implantation rates. We tested couples attending Fertility Associates, Christchurch, New Zealand for their first round of IVF. RESULTS: We found that some microbial species affected implantation. The qPCR result was interpreted qualitatively using the Z proportionality test. Samples from women at the time of Embryo Transfer who did not achieve implantation had significantly higher percent of samples that were positive for Prevotella bivia and Staphylococcus aureus compared to women who did achieve implantation. DISCUSSION: The results provide evidence that most other microbial species chosen for testing had little functional effect on implantation rates. The addition of further microbial targets (yet to be determined) could be combined in this predictive test for vaginal preparedness on the day of embryo transfer. This methodology has a substantial advantage of being affordable and easily performed in any routine molecular laboratory. This methodology is most suitable as a foundation on which to develop a timely test of microbiome profiling. Using the indicators detected to have a significant influence, these results can be extrapolated. CONCLUSION: Using a rapid antigen test, a woman can self-sample prior to embryo transfer and obtain an indication of microbial species present which could influence implantation outcome.

Collagen I dysregulation is pivotal for ovarian cancer progression.

As a principal matrisomal protein, collagen is involved in the regulation of the structural framework of extracellular matrix (ECM) and therefore is potentially crucial in determining the biophysical character of the ECM. It has been suggested that collagen architecture plays a role in ovarian cancer development, progression and therapeutic responses which led us to examine the collagen morphology in normal and cancerous ovarian tissue. Also, the behaviour of ovarian cancer cells cultured in four qualitatively different collagen gels was investigated. The results here provide evidence that collagen I morphology in the cancerous ovary is distinct from that in the normal ovary. Tumour-associated collagen I showed streams or channels of thick elongated collagen I fibrils. Moreover, fibril alignment was significantly more prevalent in endometrioid and clear cell cancers than other ovarian cancer subtypes. In this work, for the first-time collagen I architecture profiling (CAP) was introduced using histochemical staining, which distinguished between the collagen I morphologies of ovarian cancer subtypes. Immunohistochemical examination of ovarian normal and cancerous tissues also supported the notion that focal adhesion and Rho signalling are upregulated in ovarian cancers, especially in the high-grade serous tumours, as indicated by higher expression of p-FAK and p190RhoGEF. The results also support the concept that collagen I architecture, which might be collagen I concentration-dependent, influences proliferation in ovarian cancer cells. The study provides evidence that modification of collagen I architecture integrity is associated with ovarian cancer development and therapeutic responses.

Conductive Bioimprint Using Soft Lithography Technique Based on PEDOT:PSS for Biosensing.

Culture platform surface topography plays an important role in the regulation of biological cell behaviour. Understanding the mechanisms behind the roles of surface topography in cell response are central to many developments in a Lab on a Chip, medical implants and biosensors. In this work, we report on a novel development of a biocompatible conductive hydrogel (CH) made of poly (3,4-ethylenedioxythiophene):polystyrene sulfonate (PEDOT:PSS) and gelatin with bioimprinted surface features. The bioimprinted CH offers high conductivity, biocompatibility and high replication fidelity suitable for cell culture applications. The bioimprinted conductive hydrogel is developed to investigate biological cells' response to their morphological footprint and study their growth, adhesion, cell-cell interactions and proliferation as a function of conductivity. Moreover, optimization of the conductive hydrogel mixture plays an important role in achieving high imprinting resolution and conductivity. The reason behind choosing a conducive hydrogel with high resolution surface bioimprints is to improve cell monitoring while mimicking cells' natural physical environment. Bioimprints which are a 3D replication of cellular morphology have previously been shown to promote cell attachment, proliferation, differentiation and even cell response to drugs. The conductive substrate, on the other hand, enables cell impedance to be measured and monitored, which is indicative of cell viability and spread. Two dimensional profiles of the cross section of a single cell taken via Atomic Force Microscopy (AFM) from the fixed cell on glass, and its replicas on polydimethylsiloxane (PDMS) and conductive hydrogel (CH) show unprecedented replication of cellular features with an average replication fidelity of more than 90%. Furthermore, crosslinking CH films demonstrated a significant increase in electrical conductivity from 10-6 S/cm to 1 S/cm. Conductive bioimprints can provide a suitable platform for biosensing applications and potentially for monitoring implant-tissue reactions in medical devices.

Post-Covid-19 Cholangiopathy-A New Indication for Liver Transplantation: A Case Report.

Liver injury is one of the nonpulmonary manifestations described in coronavirus disease 2019 (COVID-19). Post-COVID-19 cholangiopathy is a special entity of liver injury that has been suggested as a variant of secondary sclerosing cholangitis in critically ill patients (SSC-CIP). In the general population, the outcome of SSC-CIP has been reported to be poor without orthotopic liver transplantation (OLT). However, the role of OLT for post-COVID-19 cholangiopathy is unknown. We present a case report of a 47-year-old man who recovered from acute respiratory distress syndrome from COVID-19 and subsequently developed end-stage liver disease from post-COVID-19 cholangiopathy. The patient underwent OLT and is doing well with normal liver tests for 7 months. To our knowledge, this is the first case report of a patient who underwent successful liver transplantation for post-COVID-19 cholangiopathy.

Deciphering Biophysical Modulation in Ovarian Cancer Cells.

It has been long known that the oncogenic extracellular environment plays an indispensable role in developing and nurturing cancer cell progression and in resistance to standard treatments. However, by how much the biophysical components of tumour extracellular environment contribute to these processes is uncertain. In particular, the topographic environment is scarcely explored. The biophysical modulation of cell behaviour is primarily facilitated through mechanotransduction-associated mechanisms, including focal adhesion and Rho/ROCK signalling. Dysregulation of these pathways is commonly observed in ovarian cancer and, therefore, biophysical modulation of these mechanisms may be of great importance to ovarian cancer development and progression. In this work, aspects of the biophysical environment were explored using a bioimprinting technique. The study showed that topography-mediated substrate sensing delayed cell attachment, however, cell-cell interactions overrode the effect of topography in some cell lines, such as OVCAR-5. Also, 3D topographical cues were shown to modulate the inhibition of focal adhesion and Rho signalling, which resulted in higher MAPK activity in cells on the bioimprints. It was revealed that c-Src is vital to the biophysical modulation of cell proliferation and inhibition of c-Src could downregulate biophysically modulated MAPK activity. This study provides evidence that the biophysical extracellular environment affects key intracellular mechanisms associated with tumourigenicity in ovarian cancer cells.

Lymphoepithelial cyst of the pancreas: A challenging diagnosis on fine needle aspiration.

Lymphoepithelial cysts (LECs) of the pancreas are rare, benign pancreatic cysts comprising approximately 0.5% of all pancreatic cysts. They occur predominantly in men in the 5th and 6th decades of life. LECs are true cysts lined by stratified squamous epithelium with adjacent subepithelial lymphoid tissue. They range in size from 1.2 to 17 cm (mean size 4.6 cm) and can arise in any part of the pancreas. 1 LEC resembles other benign and malignant pancreatic cysts clinically and radiologically. The cytomorphologic features of LECs have been described in a small number of case reports and it has been indicated that features may overlap with other benign and malignant pancreatic lesions. Herein, we report clinical, radiological, cytological and histopathological features of a pancreatic LEC in a 62-year-old male.

Extracellular biophysical environment: Guilty of being a modulator of drug sensitivity in ovarian cancer cells.

The roles of the extracellular biophysical environment in cancer are barely studied. This study considers the possibility that cell-like topography of a cancer cell environment may influence chemo-responses. Here, a novel bioimprinting technique was employed to produce cell-like topography on the polystyrene substrates used for cell culture. In this work, we have shown that extracellular biophysical cues generated from the topography alter the chemosensitivity of ovarian cancer cells. The three-dimensionality of the bioimprinted surface altered the cell-surface interaction, which consequently modulated intracellular signalling and chemoresponses. Sensitivity to platinum was altered more than that to paclitaxel. The effect was largely mediated through the integrin/focal adhesion system and the Rho/ROCK pathway. Moreover, the results provided evidence that biophysical cues also modulate MAPK signalling associated with chemo-resistance in ovarian cancer. Therefore, the novel findings from of this study revealed for the first time that the effects of the biophysical environment, such as substrate topography, influences ovarian cancer cell responses to clinical drugs. These observations suggest that a full clinical understanding of ovarian cancer will include biophysical aspects of tumour microenvironment.

Fatty Liver Disease: A Practical Approach.

CONTEXT.­: Fatty liver disease is now one of the most commonly encountered entities in the practice of liver pathology. Distinguishing simple steatosis from steatohepatitis is critical because the latter requires follow-up because of long-term risks that include cirrhosis and hepatocellular carcinoma. An organized approach for evaluating liver biopsies with steatosis is recommended to capture all of the relevant features: (1) degree of steatosis, (2) presence or absence of ballooning degeneration, (3) lobular inflammation, and (4) fibrosis. Herein, we provide a stepwise approach that readers can use to evaluate liver biopsies with steatosis, including examples, pitfalls, differential diagnostic considerations, and suggested diagnostic phrasing. OBJECTIVE.­: To provide a stepwise approach for the evaluation of liver biopsies showing significant steatosis (involving ≥5% of liver parenchyma). DATA SOURCES.­: Biopsies demonstrating fatty liver disease encountered in our daily practice were examined as well as recent literature. CONCLUSIONS.­: Effective evaluation of liver biopsies with steatosis requires careful histologic examination and correlation with clinical history, particularly regarding medications, nutrition status, and alcohol use. Examples of uniform reporting, including appropriate use of the nonalcoholic steatohepatitis Clinical Research Network Activity Score, are provided.

Tumour Initiation: a Discussion on Evidence for a "Load-Trigger" Mechanism.

Appropriate mechanical forces on cells are vital for normal cell behaviour and this review discusses the possibility that tumour initiation depends partly on the disruption of the normal physical architecture of the extracellular matrix (ECM) around a cell. The alterations that occur thence promote oncogene expression. Some questions, that are not answered with certainty by current consensus mechanisms of tumourigenesis, are elegantly explained by the triggering of tumours being a property of the physical characteristics of the ECM, which is operative following loading of the tumour initiation process with a relevant gene variant. Clinical observations are consistent with this alternative hypothesis which is derived from studies that have, together, accumulated an extensive variety of data incorporating biochemical, genetic and clinical findings. Thus, this review provides support for the view that the ECM may have an executive function in induction of a tumour. Overall, reported observations suggest that either restoring an ECM associated with homeostasis or targeting the related signal transduction mechanisms may possibly be utilised to modify or control the early progression of cancers. The review provides a coherent template for discussing the notion, in the context of contemporary knowledge, that tumourigenesis is an alliance of biochemistry, genetics and biophysics, in which the physical architecture of the ECM may be a fundamental component. For more definitive clarification of the concept there needs to be a phalanx of experiments conceived around direct questions that are raised by this paper.

The extracellular topographical environment influences ovarian cancer cell behaviour.

The importance of the biophysical cellular environment in cancer development has been increasingly recognised but so far has been only superficially studied. Here we investigated the effect of cell-like substrate topography on ovarian cancer cell behaviour and potential underlying signalling pathways. We observed changes in cell morphology in response to substrate topography, which implies modification of structure-function associations. Differences in focal adhesion signalling and Rho/ROCK activity suggested their involvement in the biomechanically-driven cellular responses. Cell-like topography was also shown to modulate the MAPK pathway and hence potentially regulate cell proliferation. The selective regulation of the cells by the mechanotransduction pathways that we noted has wide ranging implications for understanding cancer development. We established that the physical architecture of cell culture substrate is sufficient to influence cancer cell behaviour, independent of genetic composition or biochemical milieu.

Does the endometrial gene expression of fertile women vary within and between cycles?

STUDY QUESTION: Does gene expression of putative endometrial implantation markers vary in expression between menstrual cycles? SUMMARY ANSWER: In fertile women the expression of certain genes exhibits a pattern of stable regulation.which is not affected even when sampled twice in one cycle. WHAT IS KNOWN ALREADY: Successful implantation occurs in a minority of IVF embryo transfers. In contrast to knowledge regarding the ovulatory process, there is a sparse understanding of endometrial genes critical to implantation. This lack of knowledge hinders progress in this field. STUDY DESIGN, SIZE, DURATION: Endometrial pipelle samples were collected based on blood endocrinological markers at 2 and 7 days post initial LH surge. Five samples were collected over four cycles where the interval between collections ranged from sequential months to three years. PARTICIPANTS/MATERIALS, SETTING, METHODS: Six fertile women attending an IVF clinic for male factor infertility, had samples collected. Global gene expression profiles were obtained from laser-microdissected, endometrial glands and stroma. Nineteen potential proliferation, cytokine and adhesion markers based on previous validated reports were studied. MAIN RESULTS AND THE ROLE OF CHANCE: There was a significant modification between LH+2 and LH+7 of expression for 23 genes-11 in 8 in glands and stroma, 4 in stroma only and 3 in glands only suggesting stable, controlled regulation. Nevertheless, genes exhibited individual characteristics, e.g MKI67 exhibited lower expression at LH+7 than LH+2 and CCL4 higher, whereas TRO expressed limited difference in both cell types. Stability between cycles was demonstrated for gene expression at both LH+2-more than 60% of genes had <25% variation and at LH+7-60% had <30% variation. Further, effects of prior collection of an LH+2 sample on gene expression at LH+7 were not detected. The range of mRNA expression suggested that a clinical/diagnostic sample at LH+2 and LH+7 is likely to be a better index of endometrial function than a single sample. The possibility of redundancy suggests a panel would be more informative than a single marker. LARGE SCALE DATA: Raw and normalized microarray data have been deposited with the EMBL's European Genome-Phenome Archive for collaborative analysis, reference ega-box-815 (Lappalainen I, Almeida-King J, Kumanduri V, Senf A, Spalding JD, Ur-Rehman S, Saunders G, Kandasamy J, Caccamo M, Leinonen R et al. The European Genome-phenome Archive of human data consented for biomedical research. Nat Genet 2015;47:692-695.) [https://www.ebi.ac.uk/ega/home]. LIMITATIONS, REASONS FOR CAUTION: This type of research has difficulties of recruitment of fertile women for multiple blood testing and repeat endometrial biopsies. Therefore, these data had decreased statistical power due to the overall participant numbers. However, the inclusion of four cycles for each participant permitted the aim of obtaining information on intercycle and intracycle variability to be achieved. WIDER IMPLICATIONS OF THE FINDINGS: Our results support the feasibility of a clinical means of identification of a functional receptive endometrium. The robustness of data from individual women suggests that samples from one cycle can generally be applied to subsequent cycles. STUDY FUNDING/COMPETING INTEREST(S): Funding was granted from the Tertiary Education Commission of New Zealand, Contract I.D.:UOOX06007. There are no competing interests.

Cell-like features imprinted in the physical nano- and micro-topography of the environment modify the responses to anti-cancer drugs of endometrial cancer cells.

Topographical features of cells at nanometre resolution were fabricated in polystyrene. The study investigated the effect of physical topography on the response of cancer cells to the common anticancer drugs, paclitaxel and doxorubicin. Human endometrial cancer cells (Ishikawa) were incubated on substrates containing cell-like features that had been fabricated using our bioimprint methodology to create moulds of cells with positive (convex) and negative (concave) topography. Control cultures were performed on flat substrates. Effects of the drugs on caspase-3 expression, proliferating nuclear antigen (PCNA) expression, cell number and vascular endothelial growth factor (VEGF) secretion were determined. Results revealed that the topography influenced the cell responses in a drug-dependent manner i.e. paclitaxel effects were sensitive to topography differently to those of doxorubicin. In addition, function signalling pathways were sensitive to the detailed topography i.e. positive imprint and negative imprint induced distinct response patterns. The results in this study show for the first time that a culture surface with cell-like topography, that has both nano- and micro-resolution, influences endometrial cancer cell responses to chemotherapy drugs. The effects are dependent on the topography and also on the chemotherapy drug. In particular, the platforms described have potential to provide substrates with high physical relevancy on which to undertake preclinical testing of new drugs. The method also allows for use of different cell types to provide cell-specific topography. The results imply that physical architecture of the cancer cell environment may be a suitable prospective target to enhance clinical activity of traditional drugs. Additionally or alternatively we provide compelling support for the notion that understanding the physical component of the nano- and micro-environment may encourage a redirection of drug development. Further, our observation that the cells distinguish between the different cell-like topographies (positive and negative bioimprints) indicates that a realistic topography is advantageous as growth platforms in experiment design.

The characteristics of Ishikawa endometrial cancer cells are modified by substrate topography with cell-like features and the polymer surface.

Conventional in vitro culture studies on flat surfaces do not reproduce tissue environments, which have inherent topographical mechanical signals. To understand the impact of these mechanical signals better, we use a cell imprinting technique to replicate cell features onto hard polymer culture surfaces as an alternative platform for investigating biomechanical effects on cells; the high-resolution replication of cells offers the micro- and nanotopography experienced in typical cell-cell interactions. We call this platform a Bioimprint. Cells of an endometrial adenocarcinoma cell line, Ishikawa, were cultured on a bioimprinted substrate, in which Ishikawa cells were replicated on polymethacrylate (pMA) and polystyrene (pST), and compared to cells cultured on flat surfaces. Characteristics of cells, incorporating morphology and cell responses, including expression of adhesion-associated molecules and cell proliferation, were studied. In this project, we fabricated two different topographies for the cells to grow on: a negative imprint that creates cell-shaped hollows and a positive imprint that recreates the raised surface topography of a cell layer. We used two different substrate materials, pMA and pST. We observed that cells on imprinted substrates of both polymers, compared to cells on flat surfaces, exhibited higher expression of ß1-integrin, focal adhesion kinase, and cytokeratin-18. Compared to cells on flat surfaces, cells were larger on imprinted pMA and more in number, whereas on pST-imprinted surfaces, cells were smaller and fewer than those on a flat pST surface. This method, which provided substrates in vitro with cell-like features, enabled the study of effects of topographies that are similar to those experienced by cells in vivo. The observations establish that such a physical environment has an effect on cancer cell behavior independent of the characteristics of the substrate. The results support the concept that the physical topography of a cell's environment may modulate crucial oncological signaling pathways; this suggests the possibility of cancer therapies that target pathways associated with the response to mechanical stimuli.

A method to investigate the anti-metabolic activity of anti-cancer agents on ovarian cancer cells cultured in a 96-well high throughput format.

BACKGROUND: An early step of advanced ovarian cancer begins when floating cancerous cells as single cells or small clusters grow on the peritoneal surface. This surface is rich in extracellular matrix (ECM) proteins, which have profound effects on cellular behaviour and can facilitate cancer progression. Subsequently, this ECM may alter cellular metabolism making cancer cells susceptible to chemotherapeutic agents differently. Therefore, generating a cell culture tool in vitro that includes the interaction between ECM and cancer cells will facilitate our understanding of how cancer cells behave during cancer treatment. There is some evidence to suggest that in an in vitro model that includes ECM components such as collagens will provide a better predictive tool for drug evaluation than a traditional cell monolayer (2D) culture model. FINDINGS: As a proof -of- concept, we made a collagen gel in a 96-well plate format and utilised this to evaluate the efficacy of clinical cytotoxic drugs, a targeted drug, and food compounds in single and combination treatments. The primary endpoints were to measure the reduction of cellular metabolism and secretion of vascular endothelial growth factor (VEGF). The invasive capacity of cancer cells was observed in collagen gels and it was cell line-dependent. The responses to drugs were prominently observed in collagen gels, but they had little effect on 2D cell monolayers. These responses were cell line- and type of drug-dependent. CONCLUSIONS: The collagen gel in a 96 well plate format was easy to set up and could have potential to identify drug sensitivity in the clinical management of women with platinum resistant ovarian cancer.

Bioimprinted polymer platforms for cell culture using soft lithography.

BACKGROUND: It is becoming recognised that traditional methods of culture in vitro on flat substrates do not replicate physiological conditions well, and a number of studies have indicated that the physical environment is crucial to the directed functioning of cells in vivo. In this paper we report the development of a platform with cell-like features that is suitable for in vitro investigation of cell activity. Biological cells were imprinted in hard methacrylate copolymer using soft lithography. The cell structures were replicated at high nanometre scale resolution, as confirmed by atomic force microscopy. Optimisation of the methacrylate-based co-polymer mixture for transparency and biocompatibility was performed, and cytotoxicity and chemical stability of the cured polymer in cell culture conditions were evaluated. Cells of an endometrial adenocarcinoma cell line (Ishikawa) were cultured on bioimprinted substrates. RESULTS: The cells exhibited differential attachment on the bioimprint substrate surface compared to those on areas of flat surface and preferentially followed the pattern of the original cell footprint. CONCLUSIONS: The results revealed for the first time that the cancer cells distinguished between behavioural cues from surfaces that had features reminiscent of themselves and that of flat areas. Therefore the imprinted platform will lend itself to detailed studies of relevant physical substrate environments on cell behaviour. The material is not degraded and its permanency allows reuse of the same substrate in multiple experimental runs. It is simple and does not require expensive or specialised equipment. In this work cancer cells were studied, and the growth behaviour of the tumour-derived cells was modified by alterations of the cells' physical environment. Implications are also clear for studies in other crucial areas of health, such as wound healing and artificial tissues.

Major alteration in coxsackievirus B3 genomic RNA structure distinguishes a virulent strain from an avirulent strain.

Coxsackievirus B3 (CV-B3) is a cardiovirulent enterovirus that utilizes a 5' untranslated region (5'UTR) to complete critical viral processes. Here, we directly compared the structure of a 5'UTR from a virulent strain with that of a naturally occurring avirulent strain. Using chemical probing analysis, we identified a structural difference between the two 5'UTRs in the highly substituted stem-loop II region (SLII). For the remainder of the 5'UTR, we observed conserved structure. Comparative sequence analysis of 170 closely related enteroviruses revealed that the SLII region lacks conservation. To investigate independent folding and function, two chimeric CV-B3 strains were created by exchanging nucleotides 104-184 and repeating the 5'UTR structural analysis. Neither the parent SLII nor the remaining domains of the background 5'UTR were structurally altered by the exchange, supporting an independent mechanism of folding and function. We show that the attenuated 5'UTR lacks structure in the SLII cardiovirulence determinant.

In the secretory endometria of women, luminal epithelia exhibit gene and protein expressions that differ from those of glandular epithelia.

OBJECTIVE: To characterize the transcriptome of luminal epithelia (LE) of fertile secretory endometria and compare the results with those from glandular epithelia (GE). DESIGN: Endometrial samples were collected at 2 and 7 days after initial blood LH surge in separate menstrual cycles. LE were obtained with the use of laser microdissection. mRNA was amplified with the use of linear polymerase chain reaction and hybridized to Agilent 4×44 microarrays. Gene analysis was used to identify differentially expressed mRNAs. Immunohistochemistry was used to assess nine proteins. SETTING: One IVF clinic. PATIENT(S): Seven Caucasian fertile cycling women. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Cycle dating with the use of blood endocrinologic markers, microarrays of laser-microdissected LE, immunohistochemical analysis. RESULT(S): One hundred sixty-one (of 401) differentially expressed mRNAs in LE were identified from the metabolism pathway. Increased selective protein expression in LE at 7 days after initial LH surge was observed. LE mRNA expression was the converse of that in GE. The two cell types each had a different significant biologic pathway identified. CONCLUSION(S): Our results introduce a new concept that LE differentially expressed mRNAs are in the converse direction to that of GE, indicating different biologic processes despite the GE being continuous with the luminal monolayer. This probable distinction of biologic roles has not been noted previously. Further investigations must take cognizance of this observation.

Differences in growth properties of endometrial cancer in three dimensional (3D) culture and 2D cell monolayer.

Three-dimensional (3D) in vitro models have an invaluable role in understanding the behaviour of tumour cells in a well defined microenvironment. This is because some aspects of tumour characteristics cannot be fully recapitulated in a cell monolayer (2D). In the present study, we compared growth patterns, expression of signalling molecules, and metabolism-associated proteins of endometrial cancer cell lines in 3D and 2D cell cultures. Cancer cells formed spherical structures in 3D reconstituted basement membrane (3D rBM), and the morphological appearance was cell line dependent. Cell differentiation was observed after 8 days in the 3D rBM. There was reduced proliferation, detected by less expression of PCNA in 3D rBM than in 2D cell monolayers. The addition of exogenous epidermal growth factor (EGF) to cancer cells induced phosphorylation of EGFR and Akt in both cell culture conditions. The uptake of glucose was selectively altered in the 3D rBM, but there was a lack of association with Glut-1 expression. The secretion of vascular endothelial growth factor (VEGF) and prostaglandin E(2) (PGE(2)) was selectively altered in 3D rBM, and it was cell line dependent. Our data demonstrated that 3D rBM as an in vitro model can influence proliferation and metabolism of endometrial cancer cell behaviour compared to 2D cell monolayer. Changes are specific to individual cell types. The use of 3D rBM is, therefore, important in the in vitro study of targeted anticancer therapies.