Effects of Injectable Solutions on the Quality of Monocyte-Derived Dendritic Cells for Immunotherapy.

Therapeutic vaccines based on monocyte-derived dendritic cells have been shown to be promising strategies and may act as complementary treatments for viral infections, cancers, and, more recently, autoimmune diseases. Alpha-type-1-polarized dendritic cells (aDC1s) have been shown to induce type-1 immunity with a high capacity to produce interleukin-12p70 (IL-12p70). In the clinical use of cell-based therapeutics, injectable solutions can affect the morphology, immunophenotypic profile, and viability of cells before delivery and their survival after injection. In this sense, preparing a cell suspension that maintains the quality of aDC1s is essential to ensure effective immunotherapy. In the present study, monocytes were differentiated into aDC1s in the presence of IL-4 and GM-CSF. On day 5, the cells were matured by the addition of a cytokine cocktail consisting of IFN-α, IFN-γ, IL-1ß, TNF-α, and Poly I:C. After 48 hr, mature aDC1s were harvested and suspended in two different solutions: normal saline and Ringer's lactate. The maintenance of cells in suspension was evaluated after 4, 6, and 8 hr of storage. Cell viability, immunophenotyping, and apoptosis analyses were performed by flow cytometry. Cellular morphology was observed by electron microscopy, and the production of IL-12p70 by aDC1s was evaluated by ELISA. Compared with normal saline, Ringer's lactate solution was more effective at maintaining DC viability for up to 8 hr of incubation at 4 or 22°C.

An Insertion Within SIRPß1 Shows a Dual Effect Over Alzheimer's Disease Cognitive Decline Altering the Microglial Response.

Background: Microglial dysfunction plays a causative role in Alzheimer's disease (AD) pathogenesis. Here we focus on a germline insertion/deletion variant mapping SIRPß1, a surface receptor that triggers amyloid-ß(Aß) phagocytosis via TYROBP. Objective: To analyze the impact of this copy-number variant in SIRPß1 expression and how it affects AD molecular etiology. Methods: Copy-number variant proxy rs2209313 was evaluated in GERALD and GR@ACE longitudinal series. Hippocampal specimens of genotyped AD patients were also examined. SIRPß1 isoform-specific phagocytosis assays were performed in HEK393T cells. Results: The insertion alters the SIRPß1 protein isoform landscape compromising its ability to bind oligomeric Aß and its affinity for TYROBP. SIRPß1 Dup/Dup patients with mild cognitive impairment show an increased cerebrospinal fluid t-Tau/Aß ratio (p = 0.018) and a higher risk to develop AD (OR = 1.678, p = 0.018). MRIs showed that Dup/Dup patients exhibited a worse initial response to AD. At the moment of diagnosis, all patients showed equivalent Mini-Mental State Examination scores. However, AD patients with the duplication had less hippocampal degeneration (p < 0.001) and fewer white matter hyperintensities. In contrast, longitudinal studies indicate that patients bearing the duplication allele show a slower cognitive decline (p = 0.013). Transcriptional analysis also shows that the SIRPß1 duplication allele correlates with higher TREM2 expression and an increased microglial activation. Conclusions: The SIRPß1 internal duplication has opposite effects over MCI-to-Dementia conversion risk and AD progression, affecting microglial response to Aß. Given the pharmacological approaches focused on the TREM2-TYROBP axis, we believe that SIRPß1 structural variant might be considered as a potential modulator of this causative pathway.

Exposure to quasi-ultrafine particulate matter accelerates memory impairment and Alzheimer's disease-like neuropathology in the AppNL-G-F knock-in mouse model.

Exposure to traffic-related air pollution consisting of particulate matter (PM) is associated with cognitive decline leading to Alzheimer's disease (AD). In this study, we sought to examine the neurotoxic effects of exposure to ultrafine PM and how it exacerbates neuronal loss and AD-like neuropathology in wildtype (WT) mice and a knock-in mouse model of AD (AppNL-G-F/+-KI) when the exposure occurs at a prepathologic stage or at a later age with the presence of neuropathology. AppNL-G-F/+-KI and WT mice were exposed to concentrated ultrafine PM from local ambient air in Irvine, California, for 12 weeks, starting at 3 or 9 months of age. Particulate matter-exposed animals received concentrated ultrafine PM up to 8 times above the ambient levels, whereas control animals were exposed to purified air. Particulate matter exposure resulted in a marked impairment of memory tasks in prepathologic AppNL-G-F/+-KI mice without measurable changes in amyloid-ß pathology, synaptic degeneration, and neuroinflammation. At aged, both WT and AppNL-G-F/+-KI mice exposed to PM showed a significant memory impairment along with neuronal loss. In AppNL-G-F/+-KI mice, we also detected an increased amyloid-ß buildup and potentially harmful glial activation including ferritin-positive microglia and C3-positive astrocytes. Such glial activation could promote the cascade of degenerative consequences in the brain. Our results suggest that exposure to PM impairs cognitive function at both ages while exacerbation of AD-related pathology and neuronal loss may depend on the stage of pathology, aging, and/or state of glial activation. Further studies will be required to unveil the neurotoxic role of glial activation activated by PM exposure.

Monocyte-derived cells invade brain parenchyma and amyloid plaques in human Alzheimer's disease hippocampus.

Microglia are brain-resident myeloid cells and play a major role in the innate immune responses of the CNS and the pathogenesis of Alzheimer's disease (AD). However, the contribution of nonparenchymal or brain-infiltrated myeloid cells to disease progression remains to be demonstrated. Here, we show that monocyte-derived cells (MDC) invade brain parenchyma in advanced stages of AD continuum using transcriptional analysis and immunohistochemical characterization in post-mortem human hippocampus. Our findings demonstrated that a high proportion (60%) of demented Braak V-VI individuals was associated with up-regulation of genes rarely expressed by microglial cells and abundant in monocytes, among which stands the membrane-bound scavenger receptor for haptoglobin/hemoglobin complexes or Cd163. These Cd163-positive MDC invaded the hippocampal parenchyma, acquired a microglial-like morphology, and were located in close proximity to blood vessels. Moreover, and most interesting, these invading monocytes infiltrated the nearby amyloid plaques contributing to plaque-associated myeloid cell heterogeneity. However, in aged-matched control individuals with hippocampal amyloid pathology, no signs of MDC brain infiltration or plaque invasion were found. The previously reported microglial degeneration/dysfunction in AD hippocampus could be a key pathological factor inducing MDC recruitment. Our data suggest a clear association between MDC infiltration and endothelial activation which in turn may contribute to damage of the blood brain barrier integrity. The recruitment of monocytes could be a consequence rather than the cause of the severity of the disease. Whether monocyte infiltration is beneficial or detrimental to AD pathology remains to be fully elucidated. These findings open the opportunity to design targeted therapies, not only for microglia but also for the peripheral immune cell population to modulate amyloid pathology and provide a better understanding of the immunological mechanisms underlying the progression of AD.

Performance of standardised colposcopy to detect cervical precancer and cancer for triage of women testing positive for human papillomavirus: results from the ESTAMPA multicentric screening study.

BACKGROUND: Colposcopy, currently included in WHO recommendations as an option to triage human papillomavirus (HPV)-positive women, remains as the reference standard to guide both biopsy for confirmation of cervical precancer and cancer and treatment approaches. We aim to evaluate the performance of colposcopy to detect cervical precancer and cancer for triage in HPV-positive women. METHODS: This cross-sectional, multicentric screening study was conducted at 12 centres (including primary and secondary care centres, hospitals, laboratories, and universities) in Latin America (Argentina, Bolivia, Colombia, Costa Rica, Honduras, Mexico, Paraguay, Peru, and Uruguay). Eligible women were aged 30-64 years, sexually active, did not have a history of cervical cancer or treatment for cervical precancer or a hysterectomy, and were not planning to move outside of the study area. Women were screened with HPV DNA testing and cytology. HPV-positive women were referred to colposcopy using a standardised protocol, including biopsy collection of observed lesions, endocervical sampling for transformation zone (TZ) type 3, and treatment as needed. Women with initial normal colposcopy or no high-grade cervical lesions on histology (less than cervical intraepithelial neoplasia [CIN] grade 2) were recalled after 18 months for another HPV test to complete disease ascertainment; HPV-positive women were referred for a second colposcopy with biopsy and treatment as needed. Diagnostic accuracy of colposcopy was assessed by considering a positive test result when the colposcopic impression at the initial colposcopy was positive minor, positive major, or suspected cancer, and was considered negative otherwise. The main study outcome was histologically confirmed CIN3+ (defined as grade 3 or worse) detected at the initial visit or 18-month visit. FINDINGS: Between Dec 12, 2012, and Dec 3, 2021, 42 502 women were recruited, and 5985 (14·1%) tested positive for HPV. 4499 participants with complete disease ascertainment and follow-up were included in the analysis, with a median age of 40·6 years (IQR 34·7-49·9). CIN3+ was detected in 669 (14·9%) of 4499 women at the initial visit or 18-month visit (3530 [78·5%] negative or CIN1, 300 [6·7%] CIN2, 616 [13·7%] CIN3, and 53 [1·2%] cancers). Sensitivity was 91·2% (95% CI 88·9-93·2) for CIN3+, whereas specificity was 50·1% (48·5-51·8) for less than CIN2 and 47·1% (45·5-48·7) for less than CIN3. Sensitivity for CIN3+ significantly decreased in older women (93·5% [95% CI 91·3-95·3] in those aged 30-49 years vs 77·6% [68·6-85·0] in those aged 50-65 years; p<0·0001), whereas specificity for less than CIN2 significantly increased (45·7% [43·8-47·6] vs 61·8% [58·7-64·8]; p<0·0001). Sensitivity for CIN3+ was also significantly lower in women with negative cytology than in those with abnormal cytology (p<0·0001). INTERPRETATION: Colposcopy is accurate for CIN3+ detection in HPV-positive women. These results reflect ESTAMPA efforts in an 18-month follow-up strategy to maximise disease detection with an internationally validated clinical management protocol and regular training, including quality improvement practices. We showed that colposcopy can be optimised with proper standardisation to be used as triage in HPV-positive women. FUNDING: WHO; Pan American Health Organization; Union for International Cancer Control; National Cancer Institute (NCI); NCI Center for Global Health; National Agency for the Promotion of Research, Technological Development, and Innovation; NCI of Argentina and Colombia; Caja Costarricense de Seguro Social; National Council for Science and Technology of Paraguay; International Agency for Research on Cancer; and all local collaborative institutions.

Performance of visual inspection of the cervix with acetic acid (VIA) for triage of HPV screen-positive women: results from the ESTAMPA study.

VIA is recommended for triage of HPV-positive women attending cervical screening. In the multicentric ESTAMPA study, VIA performance for detection of cervical intraepithelial neoplasia grade 3 or worse (CIN3+) among HPV-positive women was evaluated. Women aged 30-64 years were screened with HPV testing and cytology and referred to colposcopy if either test was positive. At colposcopy visit, study-trained midwives/nurses/GPs performed VIA ahead of colposcopy. VIA was considered positive if acetowhite lesions were observed in or close to the transformation zone. Ablative treatment eligibility was assessed for VIA positives. Performance indicators were estimated. Three thousand one hundred and forty-two HPV-positive women were included. Sensitivity for CIN3+ was 85.9% (95% CI 81.2-89.5) among women <50 years and, although not significant, slightly lower in women 50+ (78.0%, 95% CI 65.9-86.6). Overall specificity was 58.6% (95% CI 56.7-60.5) and was significantly higher among women 50+ (70.3%, 95% CI 66.8-73.5) compared to women <50 (54.3%, 95% CI 52.1-56.5). VIA positivity was lower among women 50+ (35.2%, 95% CI 31.9-38.6) compared to women <50 (53.2, 95% CI 51.1-55.2). Overall eligibility for ablative treatment was 74.5% and did not differ by age. VIA sensitivity, specificity, and positivity, and ablative treatment eligibility varied highly by provider (ranges: 25%-95.4%, 44.9%-94.4%, 8.2%-65.3%, 0%-98.7%, respectively). VIA sensitivity for cervical precancer detection among HPV-positive women performed by trained providers was high with an important reduction in referral rates. However, scaling-up HPV screening triaged by VIA will be challenging due to the high variability of VIA performance and providers' need for training and supervision.

Angiotensin receptor blockade with olmesartan alleviates brain pathology in obese OLETF rats.

Metabolic syndrome (MetS) is a rapidly increasing health concern during midlife and is an emerging risk factor for the development of neurodegenerative diseases, such as Alzheimer's disease (AD). While angiotensin receptor blockers (ARB) are widely used for MetS-associated hypertension and kidney disease, its therapeutic potential in the brain during MetS are not well-described. Here, we tested whether treatment with ARB could alleviate the brain pathology and inflammation associated with MetS using the Otsuka Long-Evans Tokushima Fatty (OLETF) rat. Here, we report that chronic ARB treatment with olmesartan (10 mg/kg/day by oral gavage for 6 weeks) partially but significantly ameliorated accumulation of oxidized and ubiquitinated proteins, astrogliosis and transformation to neurotoxic astrocytes in the brain of old OLETF rats, which otherwise exhibit the progression of these pathological hallmarks associated with MetS. Additionally, olmesartan treatment restored claudin-5 and ZO-1, markers of the structural integrity of the blood-brain barrier as well as synaptic protein PSD-95, which were otherwise decreased in old OLETF rats, particularly in the hippocampus, a critical region in cognition, memory and AD. These data demonstrate that the progression of MetS in OLETF rats is associated with deterioration of various aspects of neuronal integrity that may manifest neurodegenerative conditions and that overactivation of angiotensin receptor directly or indirectly contributes to these detriments. Thus, olmesartan treatment may slow or delay the onset of degenerative process in the brain and subsequent neurological disorders associated with MetS.

Single Plane Illumination Microscopy for Microfluidic Device Imaging.

Three-dimensional imaging of live processes at a cellular level is a challenging task. It requires high-speed acquisition capabilities, low phototoxicity, and low mechanical disturbances. Three-dimensional imaging in microfluidic devices poses additional challenges as a deep penetration of the light source is required, along with a stationary setting, so the flows are not perturbed. Different types of fluorescence microscopy techniques have been used to address these limitations; particularly, confocal microscopy and light sheet fluorescence microscopy (LSFM). This manuscript proposes a novel architecture of a type of LSFM, single-plane illumination microscopy (SPIM). This custom-made microscope includes two mirror galvanometers to scan the sample vertically and reduce shadowing artifacts while avoiding unnecessary movement. In addition, two electro-tunable lenses fine-tune the focus position and reduce the scattering caused by the microfluidic devices. The microscope has been fully set up and characterized, achieving a resolution of 1.50 µm in the x-y plane and 7.93 µm in the z-direction. The proposed architecture has risen to the challenges posed when imaging microfluidic devices and live processes, as it can successfully acquire 3D volumetric images together with time-lapse recordings, and it is thus a suitable microscopic technique for live tracking miniaturized tissue and disease models.

p53 Controls Meiotic Prophase Progression and Crossover Formation.

Meiosis initiates with the formation of double strand breaks (DSBs) throughout the genome. To avoid genomic instability, these DSBs need to be correctly repaired by homologous recombination. Surveillance mechanisms involving the DNA damage response (DDR) pathway ATM-CHK2-p53 can detect the persistence of unrepaired DBSs and activate the recombination-dependent arrest at the pachytene stage. However, a complete understanding of p53 functions under normal physiological conditions remains lacking. Here, we report a detailed analysis of the p53 role during meiotic prophase in mice spermatocytes. We show that the absence of p53 regulates prophase progression by slowing down the pachytene stage when the recombination-dependent arrest occurs. Furthermore, our results show that p53 is necessary for proper crossover (CO) formation and localization. Our study contributes to a deeper understanding of p53 roles during the meiotic prophase.

Salivary, serological, and cellular immune response to the CoronaVac vaccine in health care workers with or without previous COVID-19.

We investigated the anti-SARS-CoV-2 post-vaccine response through serum and salivary antibodies, serum antibody neutralizing activity and cellular immune response in samples from health care workers who were immunized with two doses of an inactivated virus-based vaccine (CoronaVac) who had or did not have COVID-19 previously. IgA and IgG antibodies directed at the spike protein were analysed in samples of saliva and/or serum by ELISA and/or chemiluminescence assays; the neutralizing activity of serum antibodies against reference strain B, Gamma and Delta SARS-CoV-2 variants were evaluated using a virus neutralization test and SARS-CoV-2 reactive interferon-gamma T-cell were analysed by flow cytometry. CoronaVac was able to induce serum and salivary IgG anti-spike antibodies and IFN-γ producing T cells in most individuals who had recovered from COVID-19 and/or were vaccinated. Virus neutralizing activity was observed against the ancestral strain, with a reduced response against the variants. Vaccinated individuals who had previous COVID-19 presented higher responses than vaccinated individuals for all variables analysed. Our study provides evidence that the CoronaVac vaccine was able to induce the production of specific serum and saliva antibodies, serum virus neutralizing activity and cellular immune response, which were increased in previously COVID-19-infected individuals compared to uninfected individuals.

Transgenic Mouse Models of Alzheimer's Disease: An Integrative Analysis.

Alzheimer's disease (AD) constitutes the most prominent form of dementia among elderly individuals worldwide. Disease modeling using murine transgenic mice was first initiated thanks to the discovery of heritable mutations in amyloid precursor protein (APP) and presenilins (PS) genes. However, due to the repeated failure of translational applications from animal models to human patients, along with the recent advances in genetic susceptibility and our current understanding on disease biology, these models have evolved over time in an attempt to better reproduce the complexity of this devastating disease and improve their applicability. In this review, we provide a comprehensive overview about the major pathological elements of human AD (plaques, tauopathy, synaptic damage, neuronal death, neuroinflammation and glial dysfunction), discussing the knowledge that available mouse models have provided about the mechanisms underlying human disease. Moreover, we highlight the pros and cons of current models, and the revolution offered by the concomitant use of transgenic mice and omics technologies that may lead to a more rapid improvement of the present modeling battery.

SARS-CoV-2 recombinant proteins stimulate distinct cellular and humoral immune response profiles in samples from COVID-19 convalescent patients.

OBJECTIVES: In this preliminary study we investigated cellular and humoral immune responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antigens in blood samples from 14 recovered coronavirus disease 2019 (COVID-19) patients and compared them to those in samples from 12 uninfected/unvaccinated volunteers. METHODS: Cellular immunity was assessed by intracellular detection of IFN-γ in CD3+ T lymphocytes after stimulation with SARS-CoV-2 spike (S1), nucleocapsid (NC), or receptor-binding domain (RBD) recombinant proteins or overlapping peptide pools covering the sequence of SARS-CoV-2 spike, membrane and nucleocapsid regions. The humoral response was examined by ELISAs and/or chemiluminescence assays for the presence of serum IgG antibodies directed to SARS-CoV-2 proteins. RESULTS: We observed differences between humoral and cellular immune profiles in response to stimulation with the same proteins. Assays of IgG antibodies directed to SARS-CoV-2 NC, RBD and S1/S2 recombinant proteins were able to differentiate convalescent from uninfected/unvaccinated groups. Cellular immune responses to SARS-CoV-2 protein stimuli did not exhibit a specific response, as T cells from both individuals with no history of contact with SARS-CoV-2 and from recovered donors were able to produce IFN-γ. CONCLUSIONS: Determination of the cellular immune response to stimulation with a pool of SARS-CoV-2 peptides but not with SARS-CoV-2 proteins is able to distinguish convalescent individuals from unexposed individuals. Regarding the humoral immune response, the screening for serum IgG antibodies directed to SARS-CoV-2 proteins has been shown to be specific for the response of recovered individuals.

What Happens to the Immune System after Vaccination or Recovery from COVID-19?

Due to its leading role in fighting infections, the human immune system has been the focus of many studies in the context of Coronavirus disease 2019 (COVID-19). In a worldwide effort, the scientific community has transitioned from reporting about the effects of the novel coronavirus on the human body in the early days of the pandemic to exploring the body's many immunopathological and immunoprotecting properties that have improved disease treatment and enabled the development of vaccines. The aim of this review is to explain what happens to the immune system after recovery from COVID-19 and/or vaccination against SARS-CoV-2, the virus that causes the disease. We detail the way in which the immune system responds to a SARS-CoV-2 infection, including innate and adaptive measures. Then, we describe the role of vaccination, the main types of COVID-19 vaccines and how they protect us. Further, we explain the reason why immunity after COVID-19 infection plus a vaccination appears to induce a stronger response compared with virus exposure alone. Additionally, this review reports some correlates of protection from SARS-CoV-2 infection. In conclusion, we reinforce that vaccination is safe and important in achieving herd immunity.

Plaque-Associated Oligomeric Amyloid-Beta Drives Early Synaptotoxicity in APP/PS1 Mice Hippocampus: Ultrastructural Pathology Analysis.

Alzheimer's disease (AD) is a devastating neurodegenerative disorder characterized by initial memory impairments that progress to dementia. In this sense, synaptic dysfunction and loss have been established as the pathological features that best correlate with the typical early cognitive decline in this disease. At the histopathological level, post mortem AD brains typically exhibit intraneuronal neurofibrillary tangles (NFTs) along with the accumulation of amyloid-beta (Abeta) peptides in the form of extracellular deposits. Specifically, the oligomeric soluble forms of Abeta are considered the most synaptotoxic species. In addition, neuritic plaques are Abeta deposits surrounded by activated microglia and astroglia cells together with abnormal swellings of neuronal processes named dystrophic neurites. These periplaque aberrant neurites are mostly presynaptic elements and represent the first pathological indicator of synaptic dysfunction. In terms of losing synaptic proteins, the hippocampus is one of the brain regions most affected in AD patients. In this work, we report an early decline in spatial memory, along with hippocampal synaptic changes, in an amyloidogenic APP/PS1 transgenic model. Quantitative electron microscopy revealed a spatial synaptotoxic pattern around neuritic plaques with significant loss of periplaque synaptic terminals, showing rising synapse loss close to the border, especially in larger plaques. Moreover, dystrophic presynapses were filled with autophagic vesicles in detriment of the presynaptic vesicular density, probably interfering with synaptic function at very early synaptopathological disease stages. Electron immunogold labeling showed that the periphery of amyloid plaques, and the associated dystrophic neurites, was enriched in Abeta oligomers supporting an extracellular location of the synaptotoxins. Finally, the incubation of primary neurons with soluble fractions derived from 6-month-old APP/PS1 hippocampus induced significant loss of synaptic proteins, but not neuronal death. Indeed, this preclinical transgenic model could serve to investigate therapies targeted at initial stages of synaptic dysfunction relevant to the prodromal and early AD.

SARS-CoV-2 recombinant proteins stimulate distinct cellular and humoral immune response profiles in samples from COVID-19 convalescent patients

OBJECTIVES: In this preliminary study we investigated cellular and humoral immune responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antigens in blood samples from 14 recovered coronavirus disease 2019 (COVID-19) patients and compared them to those in samples from 12 uninfected/unvaccinated volunteers. METHODS: Cellular immunity was assessed by intracellular detection of IFN-γ in CD3+ T lymphocytes after stimulation with SARS-CoV-2 spike (S1), nucleocapsid (NC), or receptor-binding domain (RBD) recombinant proteins or overlapping peptide pools covering the sequence of SARS-CoV-2 spike, membrane and nucleocapsid regions. The humoral response was examined by ELISAs and/or chemiluminescence assays for the presence of serum IgG antibodies directed to SARS-CoV-2 proteins. RESULTS: We observed differences between humoral and cellular immune profiles in response to stimulation with the same proteins. Assays of IgG antibodies directed to SARS-CoV-2 NC, RBD and S1/S2 recombinant proteins were able to differentiate convalescent from uninfected/unvaccinated groups. Cellular immune responses to SARS-CoV-2 protein stimuli did not exhibit a specific response, as T cells from both individuals with no history of contact with SARS-CoV-2 and from recovered donors were able to produce IFN-γ. CONCLUSIONS: Determination of the cellular immune response to stimulation with a pool of SARS-CoV-2 peptides but not with SARS-CoV-2 proteins is able to distinguish convalescent individuals from unexposed individuals. Regarding the humoral immune response, the screening for serum IgG antibodies directed to SARS-CoV-2 proteins has been shown to be specific for the response of recovered individuals.

Human Pluripotent Stem Cell-Derived Neural Cells as a Relevant Platform for Drug Screening in Alzheimer's Disease.

Extracellular amyloid-beta deposition and intraneuronal Tau-laden neurofibrillary tangles are prime features of Alzheimer's disease (AD). The pathology of AD is very complex and still not fully understood, since different neural cell types are involved in the disease. Although neuronal function is clearly deteriorated in AD patients, recently, an increasing number of evidences have pointed towards glial cell dysfunction as one of the main causative phenomena implicated in AD pathogenesis. The complex disease pathology together with the lack of reliable disease models have precluded the development of effective therapies able to counteract disease progression. The discovery and implementation of human pluripotent stem cell technology represents an important opportunity in this field, as this system allows the generation of patient-derived cells to be used for disease modeling and therapeutic target identification and as a platform to be employed in drug discovery programs. In this review, we discuss the current studies using human pluripotent stem cells focused on AD, providing convincing evidences that this system is an excellent opportunity to advance in the comprehension of AD pathology, which will be translated to the development of the still missing effective therapies.

Multicentric study of cervical cancer screening with human papillomavirus testing and assessment of triage methods in Latin America: the ESTAMPA screening study protocol.

INTRODUCTION: Human papillomavirus (HPV) testing is replacing cytology in primary screening. Its limited specificity demands using a second (triage) test to better identify women at high-risk of cervical disease. Cytology represents the immediate triage but its low sensitivity might hamper HPV testing sensitivity, particularly in low-income and middle-income countries (LMICs), where cytology performance has been suboptimal. The ESTAMPA (EStudio multicéntrico de TAMizaje y triaje de cáncer de cuello uterino con pruebas del virus del PApiloma humano; Spanish acronym) study will: (1) evaluate the performance of different triage techniques to detect cervical precancer and (2) inform on how to implement HPV-based screening programmes in LMIC. METHODS AND ANALYSIS: Women aged 30-64 years are screened with HPV testing and Pap across 12 study centres in Latin America. Screened positives have colposcopy with biopsy and treatment of lesions. Women with no evident disease are recalled 18 months later for another HPV test; those HPV-positive undergo colposcopy with biopsy and treatment as needed. Biological specimens are collected in different visits for triage testing, which is not used for clinical management. The study outcome is histological high-grade squamous intraepithelial or worse lesions (HSIL+) under the lower anogenital squamous terminology. About 50 000 women will be screened and 500 HSIL+ cases detected (at initial and 18 months screening). Performance measures (sensitivity, specificity and predictive values) of triage techniques to detect HSIL+ will be estimated and compared with adjustment by age and study centre. ETHICS AND DISSEMINATION: The study protocol has been approved by the Ethics Committee of the International Agency for Research on Cancer (IARC), of the Pan American Health Organisation (PAHO) and by those in each participating centre. A Data and Safety Monitoring Board (DSMB) has been established to monitor progress of the study, assure participant safety, advice on scientific conduct and analysis and suggest protocol improvements. Study findings will be published in peer-reviewed journals and presented at scientific meetings. TRIAL REGISTRATION NUMBER: NCT01881659.