Effects of Gestational Intermittent Hypoxia on Placental Morphology and Fetal Development in a Murine Model of Sleep Apnea

Obstructive sleep apnea (OSA) during pregnancy is characterized by episodes of intermittent hypoxia (IH) during sleep, resulting in adverse health outcomes for mother and offspring. Despite a prevalence of 8-20% in pregnant women, this disorder is often underdiagnosed.We have developed a murine model of gestational OSA to study IH effects on pregnant mothers, placentas, fetuses, and offspring. One group of pregnant rats was exposed to IH during the last 2 weeks of gestation (GIH). One day before the delivery date, a cesarean section was performed. Other group of pregnant rats was allowed to give birth at term to study offspring's evolution.Preliminary results showed no significant weight differences in mothers and fetuses. However, the weight of GIH male offspring was significantly lower than the controls at 14 days (p < 0.01). The morphological study of the placentas showed an increase in fetal capillary branching, expansion of maternal blood spaces, and number of cells of the external trophectoderm in the tissues from GIH-exposed mothers. Additionally, the placentas from the experimental males were enlarged (p < 0.05). Further studies are needed to follow the long-term evolution of these changes to relate the histological findings of the placentas with functional development of the offspring in adulthood.Ayudas para la realización de proyectos de investigación UVa 2021 (PROYEMER 57-E.O.

Guinea Pig Oxygen-Sensing and Carotid Body Functional Properties

Mammals have developed different mechanisms to maintain oxygen supply to cells in response to hypoxia. One of those mechanisms, the carotid body (CB) chemoreceptors, is able to detect physiological hypoxia and generate homeostatic reflex responses, mainly ventilatory and cardiovascular. It has been reported that guinea pigs, originally from the Andes, have a reduced ventilatory response to hypoxia compared to other mammals, implying that CB are not completely functional, which has been related to genetically/epigenetically determined poor hypoxia-driven CB reflex. This study was performed to check the guinea pig CB response to hypoxia compared to the well-known rat hypoxic response. These experiments have explored ventilatory parameters breathing different gases mixtures, cardiovascular responses to acute hypoxia, in vitro CB response to hypoxia and other stimuli and isolated guinea pig chemoreceptor cells properties. Our findings show that guinea pigs are hypotensive and have lower arterial pO2 than rats, probably related to a low sympathetic tone and high hemoglobin affinity. Those characteristics could represent a higher tolerance to hypoxic environment than other rodents. We also find that although CB are hypo-functional not showing chronic hypoxia sensitization, a small percentage of isolated carotid body chemoreceptor cells contain tyrosine hydroxylase enzyme and voltage-dependent K+ currents and therefore can be depolarized. However hypoxia does not modify intracellular Ca2+ levels or catecholamine secretion. Guinea pigs are able to hyperventilate only in response to intense acute hypoxic stimulus, but hypercapnic response is similar to rats. Whether other brain areas are also activated by hypoxia in guinea pigs remains to be studied

Molecularly determined total tumour load in lymph nodes of stage I–II colon cancer patients correlates with high-risk factors. A multicentre prospective study

Stage I–II (pN0) colorectal cancer patients are surgically treated although up to 25 % will eventually die from disease recurrence. Lymph node (LN) status is an independent prognostic factor in colorectal cancer (CRC), and molecular tumour detection in LN of early-stage CRC patients is associated with an increased risk of disease recurrence and poor survival. This prospective multicentre study aimed to determine the relationship between LN molecular tumour burden and conventional high-risk factors in stage I–II colon cancer patients. A total of 1940 LN from 149 pathologically assessed pN0 colon cancer patients were analysed for the amount of tumour cytokeratin 19 (CK19) messenger RNA (mRNA) with the quantitative reverse transcription loop-mediated isothermal amplification molecular assay One-Step Nucleic Acid Amplification. Patient’s total tumour load (TTL) resulted from the sum of all CK19 mRNA tumour copies/μL of each positive LN from the colectomy specimen. A median of 15 LN were procured per case (IQR 12;20). Molecular positivity correlated with high-grade (p < 0.01), mucinous/signet ring type (p = 0.017), male gender (p = 0.02), number of collected LN (p = 0.012) and total LN weight per case (p < 0.01). The TTL was related to pT stage (p = 0.01) and tumour size (p < 0.01) in low-grade tumours. Multivariate logistic regression showed independent correlation of molecular positivity with gender, tumour grade and number of fresh LN [AUC = 0.71 (95 % CI = 0.62–0.79)]. Our results show that lymph node CK19 mRNA detection correlates with classical high-risk factors in stage I–II colon cancer patients. Total tumour load is a quantitative and objective measure that may help to better stage early colon cancer patients.Work supported by the Banc de Tumors-Biobanc Hospital Clinic-IDIBAPS and Xarxa de Bancs de Tumors de Catalunya (XBTC), and by grants from the Fundación Científica de la Asociación Española Contra el Cáncer (GCB13131592CAST), Ministerio de Economía y Competitividad (SAF2014–54,453-R), Agència de Gestió d’Ajuts Universitaris i de Recerca (2014SGR135), and by Sysmex Coorp Spain (Sant Just Desvern, Spain). CIBERehd is funded by the Instituto de Salud Carlos II

Estudio comparativo del control respiratorio y de la defensa frente a la Hipoxia mediados por el cuerpo carotídeo en cobaya y rata

Tesis Doctoral presentada por Elvira González Obeso para optar al grado de Doctor por la Universidad de Valladolid, Facultad de Medicina (Departamento de Bioquímica y Biología y Fisiología).-- Sujeta a Licencia Creative Commons.Se presenta un estudio compartativo morfológico y funcional del cuerpo carotídeo de cobaya, caracterizado por completo por primera vez en este trabajo de Tesis Doctoral, con el de la rata, que está bien caracterizado. El cobaya posee un cuerpo carotídeo hipotrófico comparado con el de rata. La exposición a hipoxia crónica no modificó el tamaño del cuerpo carotídeo en el cobaya pero duplicó el de rata. La cantidad de células tirosina hidroxilasa positivas en cobaya es menor que en rata (11% frente a 45% en cultivos primarios). Los parámetros ventilatorios basales en cobaya y en rata son comparables. El cobaya no hiperventila en respuesta a hipoxia aguda (10% O2) y el mismo estímulo duplica el volumen minuto en rata. En ambas especies la estimulación hipercápnica duplicó el volumen ventilatorio por minuto. Esto implica que la detección/transducción de la hipoxia a nivel del cuerpo carotídeo, o algún otro elemento del arco reflejo quimiorreceptor responsable de las respuestas ventilatorias a la hipoxia, no son funcionales en el cobaya. También se comprobó que el proceso de aclimatación a la hipoxia sostenida, que es mediado por el cuerpo carotídeo, no opera en el cobaya. Se estudió toda la cascada de trasducción del estímulo hipóxico en las células quimiorreceptoras de cobaya, comparándolas con las de rata. Se comprobó que las células quimiorreceptoras de cobaya no son activadas por la hipoxia. Sin embargo la respuesta al alto K+ extracelular demuestra que la maquinaria exocitótica de las células quimiorreceptoras del cobaya es normal. Por tanto, la incapacidad para responder a la hipoxia puede deberse a que este estímulo no es detectado por las células quimiorreceptoras de cobaya o a que algún elemento específico del acoplamiento estímulo-secrección para el estímulo hipóxico no se experesa en esta especie. El conjunto de nuestros hallazgos indican que la hipoxia no es un estímulo eficaz para las células quimiorreeptoras del cobaya, debido a que o las células no expresan el "sensor de O2" o el factor de acoplamiento entre el sensor y los canales de K+.Peer Reviewe

Effects of cigarette smoke and chronic hypoxia on airways remodeling and resistance. Clinical significance

Previously we have reported that association of cigarette smoke (CS) and chronic hypoxia (CH) interact positively to physiopathologically remodel pulmonary circulation. In present study we have exposed guinea pigs to CS smoke (four cigarettes/day; 3 months; CS) and to chronic hypoxia (12% O2, 15 days; CH) alone or in combination (CSCH animals) and evaluated airways remodeling and resistance assessed as Penh (enhance pause). We measured Penh while animals breathe air, 10% O2 and 5% CO2 and found that CS and CH animals have higher Penh than controls; Penh was even larger in CSCH animals. A rough parallelism between Penh and thickness of bronchiolar wall and muscular layer and Goblet cell number was noticed. We conclude that CS and CH association accelerates CS-induced respiratory system damage, evidenced by augmented airway resistance, bronchial wall thickness and muscularization and Goblet cell number. Our findings would suggest that appearance of hypoxia would aggravate any preexisting pulmonary pathology by increasing airways resistance and reactivity. © 2011 Elsevier B.V.The work was supported by the “Ministerio de Ciencia e Innovación of Spain”, [grant number BFU2007-61848], by the “Instituto Carlos III”, [grant number CIBER CB06/06/0050] and by “Fondo de Investigación de la Seguridad Social”, [grant number FIS 04/1424].Peer Reviewe

Guinea pig oxygen-sensing and carotid body functional properties

Mammals have developed different mechanisms to maintain oxygen supply to cells in response to hypoxia. One of those mechanisms, the carotid body (CB) chemoreceptors, is able to detect physiological hypoxia and generate homeostatic reflex responses, mainly ventilatory and cardiovascular. It has been reported that guinea pigs, originally from the Andes, have a reduced ventilatory response to hypoxia compared to other mammals, implying that CB are not completely functional, which has been related to genetically/epigenetically determined poor hypoxia-driven CB reflex. This study was performed to check the guinea pig CB response to hypoxia compared to the well-known rat hypoxic response. These experiments have explored ventilatory parameters breathing different gases mixtures, cardiovascular responses to acute hypoxia, in vitro CB response to hypoxia and other stimuli and isolated guinea pig chemoreceptor cells properties. Our findings show that guinea pigs are hypotensive and have lower arterial pO2 than rats, probably related to a low sympathetic tone and high hemoglobin affinity. Those characteristics could represent a higher tolerance to hypoxic environment than other rodents. We also find that although CB are hypo-functional not showing chronic hypoxia sensitization, a small percentage of isolated carotid body chemoreceptor cells contain tyrosine hydroxylase enzyme and voltage-dependent K+ currents and therefore can be depolarized. However hypoxia does not modify intracellular Ca2+ levels or catecholamine secretion. Guinea pigs are able to hyperventilate only in response to intense acute hypoxic stimulus, but hypercapnic response is similar to rats. Whether other brain areas are also activated by hypoxia in guinea pigs remains to be studied.This study was supported by grants BFU2015-70616R, SAF 2014-55399 and SAF 2016-77222-R (MINECO-FEDER), and CIBER CB06/06/0050 (ISCIII).We acknowledge support of the publication fee by the CSIC Open Access Support Initiative through its Unit of Information Resources for Research (URICI)Peer reviewedPeer Reviewe

Intermittent Hypoxia and Diet-Induced Obesity on the Intestinal Wall Morphology in a Murine Model of Sleep Apnea

This work analyzes the impact of two conditions, intermittent hypoxia exposure and high-fat diet in rats as models of sleep apnea. We studied the autonomic activity and histological structure of the rat jejunum and whether the overlapping of both conditions, as often observed in patients, induces more deleterious effects on the intestinal barrier. We found alterations in jejunum wall histology, predominantly in HF rats, based on increased crypt depth and submucosal thickness, as well as decreased muscularis propria thickness. These alterations were maintained with the IH and HF overlap. An increase in the number and size of goblet cells in the villi and crypts and the infiltration of eosinophils and lymphocytes in the lamina propria suggest an inflammatory status, confirmed by the increase in plasma CRP levels in all experimental groups. Regarding the CAs analysis, IH, alone or combined with HF, causes a preferential accumulation of NE in the catecholaminergic nerve fibers of the jejunum. In contrast, serotonin increases in all three experimental conditions, with the highest level in the HF group. It remains to be elucidated whether the alterations found in the present work could affect the permeability of the intestinal barrier, promoting sleep apnea-induced morbidities

Fernando de Castro and the discovery of the arterial chemoreceptors

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY).When de Castro entered the carotid body (CB) field, the organ was considered to be a small autonomic ganglion, a gland, a glomus or glomerulus, or a paraganglion. In his 1928 paper, de Castro concluded: "In sum, the Glomus caroticum is innervated by centripetal fibers, whose trophic centers are located in the sensory ganglia of the glossopharyngeal, and not by centrifugal [efferent] or secretomotor fibers as is the case for glands; these are precisely the facts which lead to suppose that the Glomus caroticum is a sensory organ." A few pages down, de Castro wrote: "The Glomus represents an organ with multiple receptors furnished with specialized receptor cells like those of other sensory organs [taste buds?]...As a plausible hypothesis we propose that the Glomus caroticum represents a sensory organ, at present the only one in its kind, dedicated to capture certain qualitative variations in the composition of blood, a function that, possibly by a reflex mechanism would have an effect on the functional activity of other organs... Therefore, the sensory fiber would not be directly stimulated by blood, but via the intermediation of the epithelial cells of the organ, which, as their structure suggests, possess a secretory function which would participate in the stimulation of the centripetal fibers." In our article we will recreate the experiments that allowed Fernando de Castro to reach this first conclusion. Also, we will scrutinize the natural endowments and the scientific knowledge that drove de Castro to make the triple hypotheses: the CB as chemoreceptor (variations in blood composition), as a secondary sensory receptor which functioning involves a chemical synapse, and as a center, origin of systemic reflexes. After a brief account of the systemic reflex effects resulting from the CB stimulation, we will complete our article with a general view of the cellular-molecular mechanisms currently thought to be involved in the functioning of this arterial chemoreceptor. © 2014 Gonzalez, Conde, Gallego-Martín, Olea, Gonzalez-Obeso, Ramirez, Yubero, Agapito, Gomez-Niño, Obeso, Rigual and Rocher.This work was supported by the Spanish Ministry of Economy and Competitiveness (Grant number BFU2012-37459 to Constancio Gonzalez), by Spanish Ministry of Health-Institute Carlos III (Grant CIBER CB06/06/0050 to Constancio Gonzalez) and by PTDC/SAU-ORG/111417/2009 (Portugal to Silvia V. Conde).Peer Reviewe

Effects of cigarette smoke and chronic hypoxia on ventilation in guinea pigs. Clinical significance

Ventilatory effects of chronic cigarette smoke (CS) alone or associated to chronic hypoxia (CH), as frequently occurs in chronic obstructive pulmonary disease (COPD), remain unknown. We have addressed this problem using whole-body plethysmography in guinea-pigs, common models to study harmful effects of CS on the respiratory system. Breathing frequencies (Bf) in control (2–5 months old) guinea pigs is 90–100 breaths/min, their tidal volume (TV) increased with age but lagged behind body weight gain and, as consequence, their minute volume (MV)/Kg decreased with age. MV did not change by acutely breathing 10% O2 but doubled while breathing 5% CO2 in air. Exposure to chronic sustained hypoxia (15 days, 12% O2, CH) did not elicit ventilatory acclimatization nor adaptation. These findings confirm the unresponsiveness of the guinea pig CB to hypoxia. Exposure to CS (3 months) increased Bf and MV but association with CH blunted CS effects. We conclude that CS and CH association accelerates CS-induced respiratory system damage leading to a hypoventilation that can worsen the ongoing COPD process.The work was supported by the “Ministerio de Ciencia e Innovación of Spain”(grant number BFU2007-61848) and by the “Instituto Carlos III”(grant number CIBER CB06/06/0050).Peer Reviewe

Un nuevo papel para el cuerpo carotídeo en la patología. Simposio en memoria del Profesor Constancio González

Resumen del trabajo presentado al XXXVIII Congreso de la Sociedad Española de Ciencias Fisiológicas (SECF), celebrado en Zaragoza del 13 al 16 de septiembre de 2016.The carotid body (CB) is a singular organ which senses variations of the arterial blood PO2, PCO2 and [H+]. In the middle of 1980s, based on our experimental data, our laboratory proposed the membrane hypothesis of acute hypoxic chemoreception formulating that the chemoreceptor cells decrease in arterial PO2 is detected by an oxygen sensor, and the reduction in the opening probability of O2-sensitive K+ channels (Lopez-Barneo et al., 1988) depolarizes chemoreceptor cells. Activation of voltage dependent Na+ and Ca2+ channels increases intracellular free [Ca2+] promoting release of catecholamines and other neurotransmitters, which augment the activity of the carotid sinus nerve producing hyperventilation (Gonzalez et al., 1994). Second messengers, such as cAMP, EPAC, and hydrogen sulphide, are capable to modulate the flow of information in the chemoreception transduction cascade (Rocher et al., 2009; Gallego-Martin et al., 2015). Nowadays, oxygen sensor identity remains unknown. Physiological systemic effects of CB activation have long been studied. In recent years, CB involvement in several pathological processes turns into a field of high interest. Some of these pathological processes are respiratory diseases that involve hypoxic situations, such as chronic sustained hypoxia in chronic obstructive pulmonary disease and chronic intermittent hypoxia (CIH) in obstructive sleep apnea (OSA). As a consequence of CIH, repetitive CB activation produces sympathetic overstimulation and redox imbalance, with high levels of reactive oxygen species (Agapito et al., 2009). CB overexcitation in OSA patients could be related to the appearance of cardiovascular pathologies (endotelial dysfunctions, hypertension, cardio- and cerebrovascular accidents), hepatometabolic alterations (obesity, insulin resistance, non-alcoholic hepatic steatosis), and neuropsychiatric diseases (anxiety, depression, dementia). Recently, it has been proposed a relationship between CIH, the main constitutive element of OSA, and cancer. Limited studies have evidenced that CIH augments growth and metastasis rate of implanted tumors in mice (Almendros et al 2013). In OSA patients, although with some discrepancies, an association between OSA and cancer incidence/mortality has been reported (Nieto et al., 2012). Discrepancies could be due to the large number of OSA-linked co-morbidities. Trying to simplify this complex pathological human situation, CIH as a single variable has been used to evaluate its effects on spontaneous tumorigenesis. In an old outbreed murine model, two intensities of CIH were applied (12% O2, moderate, and 7.5% O2, severe) mimicking two stages of OSA patients pathological situation. We have observed that long term (3 months) severe CIH augments spontaneous lung tumor incidence. These tumors are lung typical carcinoids, a type of neuroendocrine tumor. These findings could help to interpret cancer incidence in OSA patients and, on the other hand, they alert about the necessity of further designed human studies to evaluate if OSA could augment only specific types of cancer incidence.BFU2015-70616-R, MINECO/FEDER, UE; CIBERCB06/06/0050, CIBERES; APRO-I Valladolid, Asociación Española Contra el Cáncer.Peer Reviewe