Engineering mammalian cell growth dynamics for biomanufacturing.

Precise control over mammalian cell growth dynamics poses a major challenge in biopharmaceutical manufacturing. Here, we present a multi-level cell engineering strategy for the tunable regulation of growth phases in mammalian cells. Initially, we engineered mammalian death phase by employing CRISPR/Cas9 to knockout pro-apoptotic proteins Bax and Bak, resulting in a substantial attenuation of apoptosis by improving cell viability and extending culture lifespan. The second phase introduced a growth acceleration system, akin to a "gas pedal", based on an abscidic acid inducible system regulating cMYC gene expression, enabling rapid cell density increase and cell cycle control. The third phase focused on a stationary phase inducing system, comparable to a "brake pedal". A tetracycline inducible genetic circuit based on BLIMP1 gene led to cell growth cessation and arrested cell cycle upon activation. Finally, we developed a dual controllable system, combining the "gas and brake pedals", enabling for dynamic and precise orchestration of mammalian cell growth dynamics. This work exemplifies the application of synthetic biology tools and combinatorial cell engineering, offering a sophisticated framework for manipulating mammalian cell growth and providing a unique paradigm for reprogramming cell behaviour for enhancing biopharmaceutical manufacturing and further biomedical applications.

Long term culture promotes changes to growth, gene expression, and metabolism in CHO cells that are independent of production stability.

Phenotypic stability of Chinese hamster ovary (CHO) cells over long term culture (LTC) presents one of the most pressing challenges in the development of therapeutic protein manufacturing processess. However, our current understanding of the consequences of LTC on recombinant (r-) CHO cell lines is still limited, particularly as clonally-derived cell lines present distinct production stability phenotypes. This study evaluated changes of culture performance, global gene expression, and cell metabolism of two clonally-derived CHO cell lines with a stable or unstable phenotype during the LTC (early [EP] vs. late [LP] culture passages). Our findings indicated that LTC altered the behavior of CHO cells in culture, in terms of growth, overall gene expression, and cell metabolism. Regardless whether cells were categorized as stable or unstable in terms of r-protein production, CHO cells at LP presented an earlier decline in cell viability and loss of any observable stationary phase. These changes were parallelled by the upregulation of genes involved in cell proliferation and survival pathways (i.e., MAPK/ERK, PI3K-Akt). Stable and unstable CHO cell lines both showed increased consumption of glucose and amino acids at LP, with a parallel accumulation of greater amounts of lactate and TCA cycle intermediates. In terms of production stability, we found that decreased r-protein production in the unstable cell line directly correlated to the loss in r-gene copy number and r-mRNA expression. Our data revealed that LTC produced ubiquitious effects on CHO cell phenotypes, changes that were rooted in alterations in cell transcriptome and metabolome. Overall, we found that CHO cells adapted their cellular function to proliferation and survival during the LTC, some of these changes may well have limited effects on overall yield or specific productivity of the desired r-product, but they may be critical toward the capacity of cells to handle r-proteins with specific molecular features.

Engineering of Chinese hamster ovary cells for co-overexpressing MYC and XBP1s increased cell proliferation and recombinant EPO production.

Improving the cellular capacity of Chinese hamster ovary (CHO) cells to produce large amounts of therapeutic proteins remains a major challenge for the biopharmaceutical industry. In previous studies, we observed strong correlations between the performance of CHO cells and expression of two transcription factors (TFs), MYC and XBP1s. Here, we have evaluated the effective of overexpression of these two TFs on CHO cell productivity. To address this goal, we generated an EPO-producing cell line (CHOEPO) using a targeted integration approach, and subsequently engineered it to co-overexpress MYC and XBP1s (a cell line referred to as CHOCXEPO). Cells overexpressing MYC and XBP1s increased simultaneously viable cell densities and EPO production, leading to an enhanced overall performance in cultures. These improvements resulted from the individual effect of each TF in the cell behaviour (i.e., MYC-growth and XBP1s-productivity). An evaluation of the CHOCXEPO cells under different environmental conditions (temperature and media glucose concentration) indicated that CHOCXEPO cells increased cell productivity in high glucose concentration. This study showed the potential of combining TF-based cell engineering and process optimisation for increasing CHO cell productivity.

The secretory pathway - the key for unlocking the potential of Chinese hamster ovary cell factories for manufacturing therapeutic proteins.

Mammalian cell factories (in particular the CHO cell system) have been crucial in the rise of biopharmaceuticals. Mammalian cells have compartmentalized organelles where intricate networks of proteins manufacture highly sophisticated biopharmaceuticals in a specialized production pipeline - the secretory pathway. In the bioproduction context, the secretory pathway functioning is key for the effectiveness of cell factories to manufacture these life-changing medicines. This review describes the molecular components and events involved in the secretory pathway, and provides a comprehensive summary of the intracellular steps limiting the production of therapeutic proteins as well as the achievements in engineering CHO cell secretory machinery. We also consider antibody-producing plasma cells (so called "professional" secretory cells) to explore the mechanisms underpinning their unique secretory function/features. Such understandings offer the potential to further enhancement of the current CHO cell production platforms for manufacturing next generation of biopharmaceuticals.

Enhanced recombinant protein production in CHO cell continuous cultures under growth-inhibiting conditions is associated with an arrested cell cycle in G1/G0 phase.

Low temperature and sodium butyrate (NaBu) are two of the most used productivity-enhancing strategies in CHO cell cultures during biopharmaceutical manufacturing. While these two approaches alter the balance in the reciprocal relationship between cell growth and productivity, we do not fully understand their mechanisms of action beyond a gross cell growth inhibition. Here, we used continuous culture to evaluate the differential effect of low temperature and NaBu supplementation on CHO cell performance and gene expression profile. We found that an increase in cell-productivity under growth-inhibiting conditions was associated with the arrest of cells in the G1/G0 phase. A transcriptome analysis revealed that the molecular mechanisms by which low temperature and NaBu arrested cell cycle in G1/G0 differed from each other through the deregulation of different cell cycle checkpoints and regulators. The individual transcriptome changes in pattern observed in response to low temperature and NaBu were retained when these two strategies were combined, leading to an additive effect in arresting the cell cycle in G1/G0 phase. The findings presented here offer novel molecular insights about the cell cycle regulation during the CHO cell bioprocessing and its implications for increased recombinant protein production. This data provides a background for engineering productivity-enhanced CHO cell lines for continuous manufacturing.

Reprogramming of Chinese hamster ovary cells towards enhanced protein secretion.

The deficient secretory phenotype of Chinese hamster ovary (CHO) cells is a major limitation for high-level production of biopharmaceuticals, particularly for those with complex molecular architectures and post-translational modifications. To improve CHO cell secretory capacity, we recently engineered CHO cell hosts to overexpress BLIMP1 (CHOB), in a cell engineering strategy that transformed the cellular machinery and led to significantly higher product yields and cell-specific productivities for different rproteins. Here, as a follow-up to our previous study, we developed new CHO cell hosts that co-overexpress BLIMP1 and XBP1s ( CHOBX ), two transcription factors that together drive the professional secretory function of antibody-producing plasma cells. We found that the CHOBX cells presented an improved performance over that of CHOB cells, with better product yields and cell-specific productivities for a recombinant IgG1 and a 'difficult-to-express' EPO-Fc fusion protein. These improvements in the CHOBX-derived cell lines resulted from a series of physiological and metabolic changes due to the synergetic co-expression of BLIMP1 and XBP1s. Firstly, cells presented an inhibited cell growth and arrested cell cycle in G1/G0 phase, features that were directly linked to BLIMP1 expression levels. Secondly, cells increased protein translation (both overall and recombinant protein), expanded the endoplasmic reticulum and improved their capacity to secrete protein more effectively. Lastly, cells showed a metabolic profile favouring energy production, with a pronounced lactate switch and increased consumption of amino acids. This study highlights the value of transcription factors for reprogramming CHO cells towards a desired phenotype, offering the potential to engineer cells with new functionalities for enhanced manufacturing of recombinant therapeutic proteins.

Combined gene and environmental engineering offers a synergetic strategy to enhance r-protein production in Chinese hamster ovary cells.

Environmental growth-inhibition conditions (GICs) have been used extensively for increasing cell-specific productivity (qP ) of Chinese hamster ovary (CHO) cells, with the most common being temperature downshift and sodium butyrate (NaBu) treatment. B lymphocyte-induced maturation protein-1 (BLIMP1) overexpression in CHO cells can also inhibit cell growth and increase product titers and qP . Given the similar responses, this study evaluated the individual and combined effects of BLIMP1 expression, low temperature, and NaBu treatment on culture performance, cell metabolism, and recombinant protein production of CHO cells. As expected, all three interventions decreased cell growth, arrested cells in G1/G0 cell cycle phase, and increased qP . However, CHO cells presented different responses when considering cell viability, recombinant gene expression, and cell metabolism that indicated differences in the molecular loci by which BLIMP1 and GICs generated higher productivities. Combinations of BLIMP1 expression and GICs acted synergistically to inhibit cell growth and maximize r-protein production, with the BLIMP1/NaBu condition leading to the most significant improvements in product titers and qP . This latter condition also proved to substantially increase product yields (up to 9.8 g immunoglobulin G1 [IgG1]/L and 2.2 g erythropoietin-Fc [EPO-Fc]/L) and qP (up to 179 pg/cell/day [pcd] for IgG1 and 30 pcd for EPO-Fc) in high-density perfusion cultures. These findings offered mechanistic insights about the productivity-enhancing effects of BLIMP1 and GICs, as well as their complementarity for generating highly productive processes.

Overexpression of transcription factor BLIMP1/prdm1 leads to growth inhibition and enhanced secretory capacity in Chinese hamster ovary cells.

Chinese hamster ovary (CHO) cells present inherent limitations for processing and secretion of large amounts of recombinant proteins, especially for those requiring complex post-translational processing. To tackle these limitations, we engineered CHO host cells (CHOK1 and CHOS) to overexpress the transcription factor BLIMP1/prdm1 (a master regulator of the highly-secreting phenotype of antibody-producing plasma cells), generating novel CHO cell lines (referred to as CHOB). The CHOB cell lines exhibited decreased cell densities, prolonged stationary phase and arrested cell cycle in G1/G0 phase but simultaneously had significantly greater product titre for recombinant IgG1 (> 2-fold increase) coupled with a significantly greater cell-specific productivities (> 3-fold increase). We demonstrated that the improved productive phenotype of CHOB cells resulted from a series of changes to cell physiology and metabolism. CHOB cells showed a significantly greater ER size and increased protein synthesis and secretion capacity compared to control cells. In addition, CHOB cells presented a metabolic profile that favoured energy production to support increased recombinant protein production. This study indicated that a cell engineering approach based on BLIMP1 expression offers great potential for improving the secretory capacity of CHO cell hosts utilised for manufacture of recombinant biopharmaceuticals. Our findings also provides a greater understanding of the relationship between cell growth and productivity, valuable generic information for improving productive phenotypes for CHO cell lines during industrial cell line development.

Temperature Down-Shift Modifies Expression of UPR-/ERAD-Related Genes and Enhances Production of a Chimeric Fusion Protein in CHO Cells.

Low culture temperature enhances the cell-specific productivity of Chinese hamster ovary (CHO) cells expressing varied recombinant (r-) proteins, but the mechanisms remain unclear. Regulation of unfolded protein response (UPR) pathway genes, such as transcriptional regulatory factors and endoplasmic reticulum (ER)-resident proteins, appear to be involved in the improvements of r-protein production under low temperature conditions. The transcriptional regulation of UPR-specific targets is studied in response to decreased culture temperature in relation to production of a difficult-to-express protein. A clonally-derived CHO cell line expressing a chimeric fusion protein (human erythropoietin [hEPO] linked to a murine Fc region, hEPO-Fc) is evaluated in terms of growth, metabolism, r-protein production and UPR-/ER associated degradation (ERAD)-specific gene expression at standard (37 °C) and low (32 °C) temperature in batch and fed-batch systems. Low temperature decreased peak cell density, improved viability, generated cell cycle arrest in the G1 phase and enhanced hEPO-Fc expression in both batch and fed-batch cultures. A low culture temperature significantly upregulated genes encoding UPR-specific transcriptional activators (xbp1s, ddit3, and atf5) and ER-resident proteins (grp78, grp94, trib3, and ero1α), that are associated with folding and processing of proteins within the ER. Further, low culture temperature decreased expression of genes involved in ERAD (edem3, sels, herpud1, and syvn1) indicating a decreased potential for protein degradation.

Metabolic profiling of Chinese hamster ovary cell cultures at different working volumes and agitation speeds using spin tube reactors.

Culture systems based on spin tube reactors have been consolidated in the development of manufacturing processes based on Chinese hamster ovary (CHO) cells. Despite their widespread use, there is little information about the consequences of varying operational setting parameters on the culture performance of recombinant CHO cell lines. Here, we investigated the effect of varying working volumes and agitation speeds on cell growth, protein production, and cell metabolism of two clonally derived CHO cell lines (expressing an IgG1 and a "difficult-to-express" fusion protein). Interestingly, low culture volumes increased recombinant protein production and decreased cell growth, while high culture volumes had the opposite effect. Altering agitation speeds exacerbated or moderated the differences observed due to culture volume changes. Combining low agitation rates with high culture volumes suppressed growth and recombinant protein production in CHO cells. Meanwhile, high agitation rates narrowed the differences in culture performance between low and high working volumes. These differences were also reflected in cell metabolism, where low culture volumes enhanced oxidative metabolism (linked to a productive phenotype) and high culture volume generated a metabolic profile that was predominately glycolytic (linked to a proliferative phenotype). Our findings indicate that the culture volume influence on metabolism modulates the balance between cell growth and protein production, a key feature that may be useful to adjust CHO cells toward a more productive phenotype.

High glucose and low specific cell growth but not mild hypothermia improve specific r-protein productivity in chemostat culture of CHO cells.

In the biopharmaceutical sector, Chinese hamster ovary (CHO) cells have become the host of choice to produce recombinant proteins (r-proteins) due to their capacity for correct protein folding, assembly, and posttranslational modification. However, the production of therapeutic r-proteins in CHO cells is expensive and presents insufficient production yields for certain proteins. Effective culture strategies to increase productivity (qp) include a high glucose concentration in the medium and mild hypothermia (28-34 °C), but these changes lead to a reduced specific growth rate. To study the individual and combined impacts of glucose concentration, specific growth rate and mild hypothermia on culture performance and cell metabolism, we analyzed chemostat cultures of recombinant human tissue plasminogen activator (rh-tPA)-producing CHO cell lines fed with three glucose concentrations in feeding media (20, 30 and 40 mM), at two dilution rates (0.01 and 0.018 1/h) and two temperatures (33 and 37 °C). The results indicated significant changes in cell growth, cell cycle distribution, metabolism, and rh-tPA productivity in response to the varying environmental culture conditions. High glucose feed led to constrained cell growth, increased specific rh-tPA productivity and a higher number of cells in the G2/M phase. Low specific growth rate and temperature (33 °C) reduced glucose consumption and lactate production rates. Our findings indicated that a reduced specific growth rate coupled with high feed glucose significantly improves r-protein productivity in CHO cells. We also observed that low temperature significantly reduced qp, but not cell growth when dilution rate was manipulated, regardless of the glucose concentration or dilution rate. In contrast, we determined that feed glucose concentration and consumption rate were the dominant aspects of the growth and productivity in CHO cells by using multivariate analysis.

Mild hypothermia upregulates myc and xbp1s expression and improves anti-TNFα production in CHO cells.

Chinese hamster ovary (CHO) cells are the most frequently used host for commercial production of therapeutic proteins. However, their low protein productivity in culture is the main hurdle to overcome. Mild hypothermia has been established as an effective strategy to enhance protein specific productivity, although the causes of such improvement still remain unclear. The self-regulation of global transcriptional regulatory factors, such as Myc and XBP1s, seems to be involved in increased the recombinant protein production at low temperature. This study evaluated the impact of low temperature in CHO cell cultures on myc and xbp1s expression and their effects on culture performance and cell metabolism. Two anti-TNFα producing CHO cell lines were selected considering two distinct phenotypes: i.e. maximum cell growth, (CN1) and maximum specific anti-TNFα production (CN2), and cultured at 37, 33 and 31°C in a batch system. Low temperature led to an increase in the cell viability, the expression of the recombinant anti-TNFα and the production of anti-TNFα both in CN1 and CN2. The higher production of anti-TNFα in CN2 was mainly associated with the large expression of anti-TNFα. Under mild hypothermia myc and xbp1s expression levels were directly correlated to the maximal viable cell density and the specific anti-TNFα productivity, respectively. Moreover, cells showed a simultaneous metabolic shift from production to consumption of lactate and from consumption to production of glutamine, which were exacerbated by reducing culture temperature and coincided with the increased anti-TNFα production. Our current results provide new insights of the regulation of myc and xbp1s in CHO cells at low temperature, and suggest that the presence and magnitude of the metabolic shift might be a relevant metabolic marker of productive cell line.

Doppler ultrasound evaluation in preeclampsia.

BACKGROUND: Worldwide preeclampsia (PE) is the leading cause of maternal death and affects 5 to 8% of pregnant women. PE is characterized by elevated blood pressure and proteinuria. Doppler Ultrasound (US) evaluation has been considered a useful method for prediction of PE; however, there is no complete data about the most frequently altered US parameters in the pathology. The aim of this study was to evaluate the uterine, umbilical, and the middle cerebral arteries using Doppler US parameters [resistance index (RI), pulsatility index (PI), notch (N), systolic peak (SP) and their combinations] in pregnant women, in order to make a global evaluation of hemodynamic repercussion caused by the established PE. RESULTS: A total of 102 pregnant Mexican women (65 PE women and 37 normotensive women) were recruited in a cases and controls study. Blood velocity waveforms from uterine, umbilical, and middle cerebral arteries, in pregnancies from 24 to 37 weeks of gestation were recorded by trans-abdominal examination with a Toshiba Ultrasound Power Vision 6000 SSA-370A, with a 3.5 MHz convex transducer. Abnormal general Doppler US profile showed a positive association with PE [odds ratio (OR) = 2.93, 95% confidence interval (CI) = 1.2 - 7.3, P = 0.021)], and a specificity and predictive positive value of 89.2% and 88.6%, respectively. Other parameters like N presence, RI and PI of umbilical artery, as well as the PI of middle cerebral artery, showed differences between groups (P values < 0.05). CONCLUSION: General Doppler US result, as well as N from uterine vessel, RI from umbilical artery, and PI from umbilical and middle cerebral arteries in their individual form, may be considered as tools to determine hemodynamic repercussion caused by PE.

Diagnóstico prenatal de cierre prematuro del foramen oval e hidrops fetal / Prenatal diagnosis of foramen ovale premature closure and hydrops fetalis

A case of premature closure of foramen ovale (PCFO) is presented. The patient was sent to the Fetal Maternal Unit after 30 weeks of gestation due to hydrops fetalis. A Doppler ultrasound performed after 34 weeks of gestation showed no interatrial flow and led to the confirmation of PCFO, associated to pleural effusion and ascitis. No evidence of hydrops, pleural effusion, ascitis, cardiac failure, cardiac defects nor chromosomal abnormalities were present in the newborn baby. Since prenatal diagnosis of PCFO is a life threatening condition, detection improves fetal and neonatal life expectancy (AU)

Se reporta el cierre prematuro de foramen oval (CPFO) en un embarazo de 30 semanas de gestación, enviado con hidrops fetal a Unidad de Medicina Materno Fetal. A las 34 semanas se evaluó con Ultrasonido Doppler y se observó falta de flujo entre aurículas, confirmando el CPFO, presencia de derrame pleural y ascitis. Después del nacimiento, al neonato se le descartó la presencia de hidrops, derrame o ascitis, insuficiencia cardiaca, defectos cardiacos o cromosomopatía, y egresó sano. El CPFO pone en riesgo la vida. Cuando es detectado prenatalmente mejora la expectativa de vida fetal y neonatal (AU)

[Pituitary tumor detected prenatally]. / Tumor hipofisiario detectado prenatalmente.

INTRODUCTION: Brain tumors are present in 2.9 per 100,000 newborn. Craniopharyngioma is a benign and slow growing brain tumor, frequently localized in the sellar and suprasellar region. There are few reports of pituitary tumor detected prenatally. CASE REPORT: We report a neonate with a craniopharyngioma detected prenatally as a pituitary tumor. In a 23 year old mother, second gestation, with no important history, was detected a sellar tumor at 31 gestation weeks, the obstetric ultrasound reported a suprasellar tumor of 2 per 3 cm diameter. Pregnancy ended in a vaginal delivery at 39 weeks, and obtained a 3.9 kg female, with cephalic diameter of 37.5 cm, the Apgar score was 8-9 at 1st and 5th minutes. In early neonatal period was scanned and confirmed a 3.2/2.3/2.9 cm suprasellar tumor with calcium deposits. The Paediatric Oncology department suggested a surgery and was realized a craniotomy at 3rd week of age. The surgery allowed to obtain 30% of the tumor and confirmed by histology craniopharyngioma. Patient had favourable evolution and was discharged at 3 months of age. CONCLUSIONS: We report a neonate in who was detected by prenatal ultrasound the presence of a suprasellar solid tumor, scan and magnetic resonance images in neonatal period defined its size and location and a craniopharyngioma was confirmed by histology. Patient had a satisfactory postsurgical evolution and was discharged at 3 months of age.

[Prenatal diagnosis of pulmonary valve stenosis and immediate neonal valvuloplasty]. / Diagnóstico prenatal de estenosis pulmonar grave y valvuloplastia neonatal inmediata. Reporte de un caso.

UNLABELLED: There are few reports of prenatal diagnosis of severe pulmonary valvar stenosis (PVS). It affects 1/22,000 newborn and represents 8-10% of total congenital cardiac defects. Clinic CASE: we report a case of a neonate in which was prenatally detected a pulmonary valvar stenosis and was successfully corrected with early valvuloplasty. From a 36-Year-old woman sent to evaluation to the fetal maternal unit because a tricuspid valvar insufficiency detected at 36 gestation weeks (GW). A VPS was suspected before born and a pregnancy ended in programated caesarean delivery at 38 GW, obtaining a 3 kg male, in which early echocardiography reported a severe PVS, promptly was initiated prostaglandin E1 (PgE1) infusion avoiding patent ductus arteriosus (PDA) closure, following a percutaneus balloon dilatation valvuloplasty at 48 hours, improving cyanosis and transvalvular Doppler flow. CONCLUSION: we report a neonate referred with an opportune prenatal diagnosis of tricuspid insufficiency and confirmed a severe PVS, PgE1 was infused immediately after born, allowing successfully balloon dilatation valvuloplasty in first 48 hours.

Amniotic fluid index accuracy in mid-trimester.

OBJECTIVE: The aim of this trial was to evaluate the correlation between amniotic fluid index (AFI) and the real amniotic volume in mid-trimester pregnancies. METHODS: Eight women with mid-trimester anhydramnios pregnancies were included. Patients gave their informed consent. Those with premature rupture of membranes were excluded. Amnioinfusion was performed by instilling up to 400 ml of saline solution in 100 ml aliquots. With the patient in supine position, the AFI was determined by the sum of measurements of the deepest vertical pools in each of the 4 quadrants of the maternal uterus, evaluating the AFI after every 100 ml of infused solution. Statistical analysis was performed by analysis of variance, determination coefficient, linear correlation and t test. RESULTS: We realized 29 AFI measurements after amnioinfusions of 100, 200, 300 and 400 ml of saline solution. The AFI ranges that corresponded to the infused volume were: after 100 ml = 5-11 cm (median 6.5 cm); after 200 ml = 9-12.7 cm (median 10.3 cm); after 300 ml = 10-17 cm (median 13.5 cm), and after 400 ml = 12-16 cm (median 15 cm). The correlation between the AFI and the volume infused was r = 0.81 (p < 0.0001), the variance was r(2) = 0.65, suggesting that 65% of the variation in AFI measurements is directly accounted for by the amniotic fluid volume and 35% of the AFI measurements is accounted for by factors other than amniotic volume. CONCLUSIONS: We obtained a strong correlation between AFI and amnioinfusion volume (median of 6.5, 10.3, 13.5 and 15 cm after amniotic infusion of 100, 200, 300 and 400 ml of saline solution, respectively). The variance (0.65) suggests that 35% of AFI measurements is accounted for by factors other than amniotic fluid volume.

[Prenatal diagnosis of polycystic kidneys and Mullerian duplication. A case report]. / Diagnóstico prenatal de riñones poliquísticos y duplicación mulleriana. Reporte de un caso.

Polycystic kidney disease is a common genetic cause of chronic kidney disease, characterized by the formation of multiple cysts in the kidneys and other organs, occurs in 1 in 20,000 live births. 30 to 50% of affected newborns die shortly after birth because of respiratory and renal insufficiency. This study reports the case of a newborn with polycystic kidney disease diagnosed by obstetric ultrasound at 26 weeks of gestation and kidneys anhidramnios due to increased volume and appearance "in sponge." Neonato a primigravida 19 years of age. At 26 weeks you will be detected in a routine obstetric evaluation that measures were in line for somatométricas fetal gestational age, and anhidramnios increase in renal mass and bilateral thorax narrow. The pregnancy ended in a cesarean section at 37 weeks with a newborn of 3140 g, women who died within minutes after birth. We requested an autopsy because of the need for genetic counseling. The findings were: enlarged kidneys with microcystic dilatation of collecting duct and in the renal cortex and medulla, liver fibrosis and Müllerian duplication with double uterine cavity. It is a rare association between polycystic kidney disease and Müllerian duplication, liver fibrosis confirmed by autopsy and has not been documented previously.

[Multiple pregnancies prevalence: its raise on last decade]. / Prevalencia de embarazos múltiples: incremento en la ultima década.

BACKGROUND: Multiple pregnancies prevalence has been increasing in last decade, which have also increased the requirements of neonatal intensive care units and all problems related to premature neonate or low birth weight. Prevalence rate of twin (18 to 26 in 1,000 births), and triple pregnancies (0.37 to 1.74 in 1,000 births) have raised too, perhaps due to assisted reproductive techniques. OBJECTIVE: To know incidence of multiple pregnancies at Unidad Medica de Alta Especialidad no. 23, from Institute Mexicano del Seguro Social. MATERIAL AND METHOD: Retrospective and descriptive study. We review the files of multiple pregnancies from 1972 to 2006 to estimate its rate and change every five and ten years. RESULTS: We registered 9,055 twin pregnancies during the period, with a rate of 7.1 to 14.4 in 1,000 (63% of increase in the last decade [12.6 in 1,000 births] compared with the previous decade [7.7 in 1,000 births]; p < 0.005). Pregnancies with three or more fetuses were 202, with 191 triplets, 13 with four, three with five, and one with six products (646 newborns). Incidence of multiple pregnancies with four or more products has also increased in last decade: 230 times higher than two decades before. CONCLUSION: Multiple pregnancies rate has increased in last decade: 63% in twin pregnancies, 217% in triplets, and 230 times more than expected in four or more products pregnancies.

[Usefulness of Doppler ultrasonography to detect fetal anemia in rh alloimmunized pregnancies]. / Ultrasonido Doppler para la detección de anemia fetal por isoimmunización a Rh.

BACKGROUND: pregnant women with Rh alloimmunization (RhA) are submitted to invasive procedures to assess fetal anemia (FA). Recently a non-invasive approach to FA diagnosis has been proposed using Doppler ultrasound (DU) to identify increased peak velocity of systolic blood flow (Vm) in the middle cerebral artery (MCA). METHODOLOGY: eleven Rh alloimmunized pregnant women with serum red-cell antibody titers > 1:16 were included. Twenty-four procedures were done measuring the VmMCA followed by cordocentesis and fetal hemoglobin (FH) analysis. Pearson's linear correlation was calculated between the multiples of the median (MoM) of the VmMCA and the MoM of the FH, as well as the sensitivity, specificity and positive predictive value (PPV) for FA prediction. RESULTS: we found FA (FH mean = 6 g/dL) in 12 of 24 evaluations with a VmMCA mean of 1.5 MoM and a range from 1.22 to 1.68 MoM; in the remaining 12 cases the FH was normal (FH mean = 13.1 g/dL) with a VmMCA mean of 0.97 MoM and a range from 0.35-1.17 MoM (p < 0.001). Eleven fetuses with anemia had a MoM of the VmMCA above 1.29, except one with 1.22 MoM. The linear correlation between the MoM of the VmMCA and the MoM of FH was 0.83. The sensitivity of the MoM of the VmMCA to detect FA was 91%, specificity of 100% and PPV of 100%. CONCLUSIONS: DU measurement of the VmMCA was a useful non-invasive technique to evaluate FA. The sensitivity and PPV for FA diagnosis in RhA was above 90%.