Impact of α-synuclein pathology on transplanted hESC-derived dopaminergic neurons in a humanized α-synuclein rat model of PD.

Preclinical assessment of the therapeutic potential of dopamine (DA) neuron replacement in Parkinson's disease (PD) has primarily been performed in the 6-hydroxydopamine toxin model. While this is a good model to assess graft function, it does not reflect the pathological features or progressive nature of the disease. In this study, we establish a humanized transplantation model of PD that better recapitulates the main disease features, obtained by coinjection of preformed human α-synuclein (α-syn) fibrils and adeno-associated virus (AAV) expressing human wild-type α-syn unilaterally into the rat substantia nigra (SN). This model gives rise to DA neuron dysfunction and progressive loss of DA neurons from the SN and terminals in the striatum, accompanied by extensive α-syn pathology and a prominent inflammatory response, making it an interesting and relevant model in which to examine long-term function and integrity of transplanted neurons in a PD-like brain. We transplanted DA neurons derived from human embryonic stem cells (hESCs) into the striatum and assessed their survival, growth, and function over 6 to 18 wk. We show that the transplanted cells, even in the presence of ongoing pathology, are capable of innervating the DA-depleted striatum. However, on closer examination of the grafts, we found evidence of α-syn pathology in the form of inclusions of phosphorylated α-syn in a small fraction of the grafted DA neurons, indicating host-to-graft transfer of α-syn pathology, a phenomenon that has previously been observed in PD patients receiving fetal tissue grafts but has not been possible to demonstrate and study in toxin-based animal models.

A systematic capsid evolution approach performed in vivo for the design of AAV vectors with tailored properties and tropism.

Adeno-associated virus (AAV) capsid modification enables the generation of recombinant vectors with tailored properties and tropism. Most approaches to date depend on random screening, enrichment, and serendipity. The approach explored here, called BRAVE (barcoded rational AAV vector evolution), enables efficient selection of engineered capsid structures on a large scale using only a single screening round in vivo. The approach stands in contrast to previous methods that require multiple generations of enrichment. With the BRAVE approach, each virus particle displays a peptide, derived from a protein, of known function on the AAV capsid surface, and a unique molecular barcode in the packaged genome. The sequencing of RNA-expressed barcodes from a single-generation in vivo screen allows the mapping of putative binding sequences from hundreds of proteins simultaneously. Using the BRAVE approach and hidden Markov model-based clustering, we present 25 synthetic capsid variants with refined properties, such as retrograde axonal transport in specific subtypes of neurons, as shown for both rodent and human dopaminergic neurons.

Tr-1-like CD4+CD25-CD127-/lowFOXP3- cells are the main source of interleukin 10 in patients with cutaneous leishmaniasis due to Leishmania braziliensis.

CD4(+)CD25(+)FOXP3(+) regulatory T cells have long been shown to mediate susceptibility to Leishmania infection, mainly via interleukin 10 production. In this work, we showed that the main sources of interleukin 10 in peripheral blood mononuclear cells (PBMCs) from patients with cutaneous leishmaniasis due to Leishmania braziliensis are CD4(+)CD25(-)CD127(-/low)FOXP3(-) cells. Compared with uninfected controls, patients with CL had increased frequencies of circulating interleukin 10-producing CD4(+)CD25(-)CD127(-/low) cells, which efficiently suppressed tumor necrosis factor α production by the total PBMC population. Also, in CL lesions, interleukin 10 was mainly produced by CD4(+)CD25(-) cells, and interleukin 10 messenger RNA expression was associated with interleukin 27, interleukin 21, and interferon γ expression, rather than with FOXP3 or transforming growth factor ß expressions. Active production of both interleukin 27 and interleukin 21, together with production of interferon γ and interleukin 10, was also detected in the lesions. Since these cytokines are associated with the differentiation and activity of Tr-1 cells, our results suggest that this cell population may play an important role in the immunomodulation of CL. Therefore, development of treatments that interfere with this pathway may lead to faster parasite elimination.

Endoscopy-assisted laparoscopic wedge-resection of gastric glomus tumor: A case report.

INTRODUCTION: Glomus tumor is a pericytic mesenchymal neoplasm that most commonly occurs in the extremities. The occurrence in visceral organs is rare and is a differential diagnosis with other gastric submucosal tumors. PRESENTATION OF CASE: A woman with epigastric pain underwent esophagogastroduodenoscopy (EGD) which revealed a gastric submucosal tumor. Endoscopic ultrasound with fine-needle aspiration allowed preoperative diagnosis of gastric glomus tumor. Intraoperative EGD-assisted laparoscopic segmental gastrectomy was successfully performed. The patient was discharged in the second postoperative day. There was no evidence of recurrence at 8 months of follow-up. DISCUSSION: The stomach is a rare location for the glomus tumor, a neoplasm of the glomus body, which is a perivascular structure with thermoregulatory function. Preoperative diagnosis is challenging, and endoscopic ultrasound (EUS) is useful for both assessing malignancy-associated features and biopsy guiding. The treatment is surgical resection with attention to adequate oncological margins while preserving healthy gastric wall. CONCLUSION: Immunohistochemical analysis of specimen obtained by EUS fine-needle allows accurate preoperative diagnosis and laparoscopic-endoscopic combined surgery allows good oncological and functional results.

Exploring wine yeast natural biodiversity to select strains with enological traits adapted to climate change.

Wine is widely consumed throughout the world and represents a significant financial market, but production faces increasing challenges. While consumers progressively value more complex flavor profiles, regional authenticity, and decreased use of additives, winemakers strive for consistency among climate change, characterized by rising environmental temperatures and sun burn events. This often leads to grapes reaching phenolic maturity with higher sugar levels, and increased microbial spoilage risk. Herein, we addressed these dual concerns by investigating the use of autochthonous Saccharomyces cerevisiae strains for fermentations of grape musts resulting from these altered conditions. We characterized underexplored repositories of naturally-occurring strains isolated from different environments and geographical regions, regarding adequate enological properties (e.g., high cell growth, reduced production of H2S, ethanol and acetic acid, increased SO2 resistance, killer activity), and other less frequently investigated properties (resistance to osmotic stress, potassium and aluminium silicates and fungicides). The phenotypic data were organized in a biobank, and bioinformatic analysis grouped the strains according to their characteristics. Furthermore, we analyzed the potential of four Portuguese isolates to be used in fermentations of grape musts with high sugar levels, uncovering promising candidates. This research therefore contributes to ongoing efforts to increase sustainability and quality of wine production.

Room temperature storage of myrtle (Eugenia gracillima Kiaersk.) tropical juice: Effects of physical and chemical preservation methods.

Tropical fruit juices produced from native fruits have been widely marketed by small agribusinesses in the Brazilian semiarid region, necessitating a deeper understanding of the impact of preservation methods on quality parameters. This study aimed to prepare myrtle (Eugenia gracillima Kiaersk.) tropical juice and investigate the effects of physical preservation (90 °C for 60 s) and chemical preservation (potassium sorbate and sodium benzoate) methods. Tropical juice formulations were evaluated after preparation and every 15 days during 60 days of storage in high-density polyethylene bottles at room temperature (25 ± 2 °C). Microbiological parameters, optical microscopy, physicochemical and bioactive parameters, antioxidant capacity, and color parameters were determined. Heat-treated tropical juice showed low counts of all microbiological parameters, but optical microscopy revealed the presence of filamentous fungi after 60 days of storage. Combined use of potassium sorbate and sodium benzoate effectively prevented the development of total yeasts and molds up to 28 days of storage. Bioactive compounds in myrtle pulp contribute to storage stability, mainly total phenolics, estimated at 855.86 mg gallic acid equivalents 100 g-1. The results suggest that it is possible to harness the economic and agroindustrial potential of E. gracillima Kiaersk. fruits for the production of tropical juices, but it is recommended that other technologies be explored, such as aseptic processing or the combined use of physical and chemical methods.

Examination of Raw Samples and Ethanol Extracts of Gerês Propolis Collected in Different Years.

Propolis, a natural resin created by bees, has garnered significant attention from both the scientific community and industry due to an impressive range of bioactivities. Nonetheless, the intrinsic variability in its chemical composition and bioactive profiles has been hindering propolis' full potential use. We previously showed that ethanol extracts (EEs) of a Portuguese propolis sample (Gerês) collected over four consecutive years displayed similar chemical and biological profiles, a constancy never documented before. However, the characteristics of the unprocessed samples of Gerês propolis were never described. Hence, the central objective of this study is to assess the quality parameters of unprocessed propolis samples collected from Gerês (G), over a four-year period (2019-2022), alongside the analysis of the chemical composition and bioactivities of the EEs prepared with the same raw samples. The ash, wax, balsam and water contents of the unprocessed samples-G19 to G22-showed minor fluctuations, likely attributed to uncontrollable natural events impacting the propolis source and collection process. On the other hand, the antimicrobial and antioxidant activities of all the four ethanol extracts (G19.EE-G22.EE) consistently align with prior studies. Furthermore, the Gerês propolis extracts showed remarkable uniformity in chemical composition parameters too, particularly concerning total polyphenol, flavonoid and ortho-diphenol contents. In summary, our research reinforces the beneficial properties of propolis and show that extracts' bioactivities remain within the reference ranges for Gerês propolis, despite minor differences in unprocessed samples, suggesting a consistent action over time. Thus, this work could be instrumental towards the establishment of standard parameters for propolis applications, offering valuable insights to this field of propolis research.

Deconvolution of spatial sequencing provides accurate characterization of hESC-derived DA transplants in vivo.

Cell therapy for Parkinson's disease has experienced substantial growth in the past decades with several ongoing clinical trials. Despite increasing refinement of differentiation protocols and standardization of the transplanted neural precursors, the transcriptomic analysis of cells in the transplant after its full maturation in vivo has not been thoroughly investigated. Here, we present spatial transcriptomics analysis of fully differentiated grafts in their host tissue. Unlike earlier transcriptomics analyses using single-cell technologies, we observe that cells derived from human embryonic stem cells (hESCs) in the grafts adopt mature dopaminergic signatures. We show that the presence of phenotypic dopaminergic genes, which were found to be differentially expressed in the transplants, is concentrated toward the edges of the grafts, in agreement with the immunohistochemical analyses. Deconvolution shows dopamine neurons being the dominating cell type in many features beneath the graft area. These findings further support the preferred environmental niche of TH-positive cells and confirm their dopaminergic phenotype through the presence of multiple dopaminergic markers.

Preclinical quality, safety, and efficacy of a human embryonic stem cell-derived product for the treatment of Parkinson's disease, STEM-PD.

Cell replacement therapies for Parkinson's disease (PD) based on transplantation of pluripotent stem cell-derived dopaminergic neurons are now entering clinical trials. Here, we present quality, safety, and efficacy data supporting the first-in-human STEM-PD phase I/IIa clinical trial along with the trial design. The STEM-PD product was manufactured under GMP and quality tested in vitro and in vivo to meet regulatory requirements. Importantly, no adverse effects were observed upon testing of the product in a 39-week rat GLP safety study for toxicity, tumorigenicity, and biodistribution, and a non-GLP efficacy study confirmed that the transplanted cells mediated full functional recovery in a pre-clinical rat model of PD. We further observed highly comparable efficacy results between two different GMP batches, verifying that the product can be serially manufactured. A fully in vivo-tested batch of STEM-PD is now being used in a clinical trial of 8 patients with moderate PD, initiated in 2022.

A novel two-factor monosynaptic TRIO tracing method for assessment of circuit integration of hESC-derived dopamine transplants.

Transplantation in Parkinson's disease using human embryonic stem cell (hESC)-derived dopaminergic (DA) neurons is a promising future treatment option. However, many of the mechanisms that govern their differentiation, maturation, and integration into the host circuitry remain elusive. Here, we engrafted hESCs differentiated toward a ventral midbrain DA phenotype into the midbrain of a preclinical rodent model of Parkinson's disease. We then injected a novel DA-neurotropic retrograde MNM008 adeno-associated virus vector capsid, into specific DA target regions to generate starter cells based on their axonal projections. Using monosynaptic rabies-based tracing, we demonstrated for the first time that grafted hESC-derived DA neurons receive distinctly different afferent inputs depending on their projections. The similarities to the host DA system suggest a previously unknown directed circuit integration. By evaluating the differential host-to-graft connectivity based on projection patterns, this novel approach offers a tool to answer outstanding questions regarding the integration of grafted hESC-derived DA neurons.

Gastric metastasis from breast cancer in male patient - Case report and literature review.

INTRODUCTION: Male breast cancer has a low incidence, and its metastases are considered even rarer and a sign of seriousness for the patient. Breast metastases are difficult to diagnose even in female patients, with few cases of gastric metastasis described in the current literature. PRESENTATION OF CASE: Male patient, 54 years old, obese, diagnosed with breast cancer for 8 years, underwent 5 years of cancer follow-up. After this period, he began to experience epigastric pain and weight loss, among other symptoms. Endoscopy was performed, showing rounded elevated lesions with central depression. After anatomopathological and biochemical tests, a diagnosis of differentiated carcinoma as a metastasis of breast origin was concluded. DISCUSSION: MBC is rare when compared to the incidence in females. Its risk factors differ between genders, and due to its rarity, there are few studies in the male population. This fact makes the evaluation of metastases more difficult. Gastric metastasis has a very low incidence even in women, approximately 8.9% of all breast cancer metastasis's locations. Other pathologies can mimic GM, always associating anatomopathological and biochemical tests. Oncological follow-up must always be carried out, and the possibility of screening must be evaluated. CONCLUSION: Despite the rare incidence and little information in the literature, the association between MBC and GM should be considered. Currently, there is no guidance for conducting systematic screenings and treatment is usually palliative due to the severity of the disease.

Toward Generating Subtype-Specific Mesencephalic Dopaminergic Neurons in vitro.

Mesencephalic dopaminergic (mDA) neurons derived from pluripotent stem cells (PSCs) have proven to be pivotal for disease modeling studies and as a source of transplantable tissue for regenerative therapies in Parkinson's disease (PD). Current differentiation protocols can generate standardized and reproducible cell products of dopaminergic neurons that elicit the characteristic transcriptional profile of ventral midbrain. Nonetheless, dopamine neurons residing in the mesencephalon comprise distinct groups of cells within diffusely defined anatomical boundaries and with distinct functional, electrophysiological, and molecular properties. Here we review recent single cell sequencing studies that are shedding light on the neuronal heterogeneity within the mesencephalon and discuss how resolving the complex molecular profile of distinct sub-populations within this region could help refine patterning and quality control assessment of PSC-derived mDA neurons to subtype-specificity in vitro. In turn, such advances would have important impact in improving cell replacement therapy, disease mechanistic studies and drug screening in PD.

Translation and validation of the addiction-like Eating Behavior Scale from English to Portuguese in Brazil.

Eating addiction is an important condition related to obesity. The Addiction-like Eating Behavior Scale (AEBS) was recently validated in English to evaluate addictive eating behaviors. To perform the translation, cultural adaptation, and validation of the AEBS scale to Portuguese to identify eating addictions in the Brazilian population. The AEBS translation, adaptation, and validation followed previously published criteria, including the steps of translation, back translation, evaluation by a committee of judges, and a pretest with 40 students. Subsequently, 150 students from the Medical School at the Recife Campus of the Federal University of Pernambuco were tested using the AEBS-Portuguese version (AEBS-p). Of these, 37 were evaluated a second time for a test-retest analysis. For data collection, 5 instruments were used: the AEBS-p, a modified version of the Yale Food Addiction Scale 2.0 (mYFAS 2.0)-Portuguese version, the Patient Health Questionnaire-9, the Binge Eating Scale (BES), and the Socio-demographic Questionnaire. The AEBS-p showed good reliability, based on a significant internal consistency value (Cronbach's alpha = 0.91), and good correlation in the test-retest evaluation. A committee of judges considered the content validity appropriate. Convergent validation was positive with the mYFAS 2.0 and BES (p < 0.001). The AEBS-p showed a significant correlation with body mass index (BMI) (p < 0.001) and significantly predicted variance in BMI that was accounted for by both the mYFAS 2.0 and BES (p = 0.022). The AEBS-p demonstrated good reliability and validity for evaluating addictive eating behaviors in the Brazilian population.

Single cell transcriptomics identifies stem cell-derived graft composition in a model of Parkinson's disease.

Cell replacement is a long-standing and realistic goal for the treatment of Parkinson's disease (PD). Cells for transplantation can be obtained from fetal brain tissue or from stem cells. However, after transplantation, dopamine (DA) neurons are seen to be a minor component of grafts, and it has remained difficult to determine the identity of other cell types. Here, we report analysis by single-cell RNA sequencing (scRNA-seq) combined with comprehensive histological analyses to characterize intracerebral grafts from human embryonic stem cells (hESCs) and fetal tissue after functional maturation in a pre-clinical rat PD model. We show that neurons and astrocytes are major components in both fetal and stem cell-derived grafts. Additionally, we identify a cell type closely resembling a class of recently identified perivascular-like cells in stem cell-derived grafts. Thus, this study uncovers previously unknown cellular diversity in a clinically relevant cell replacement PD model.

Author Correction: Single cell transcriptomics identifies stem cell-derived graft composition in a model of Parkinson's disease.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

hESC-Derived Dopaminergic Transplants Integrate into Basal Ganglia Circuitry in a Preclinical Model of Parkinson's Disease.

Cell replacement is currently being explored as a therapeutic approach for neurodegenerative disease. Using stem cells as a source, transplantable progenitors can now be generated under conditions compliant with clinical application in patients. In this study, we elucidate factors controlling target-appropriate innervation and circuitry integration of human embryonic stem cell (hESC)-derived grafts after transplantation to the adult brain. We show that cell-intrinsic factors determine graft-derived axonal innervation, whereas synaptic inputs from host neurons primarily reflect the graft location. Furthermore, we provide evidence that hESC-derived dopaminergic grafts transplanted in a long-term preclinical rat model of Parkinson's disease (PD) receive synaptic input from subtypes of host cortical, striatal, and pallidal neurons that are known to regulate the function of endogenous nigral dopamine neurons. This refined understanding of how graft neurons integrate with host circuitry will be important for the design of clinical stem-cell-based replacement therapies for PD, as well as for other neurodegenerative diseases.

Bowel obstruction in entero-colic fistula associated with magnet ingestion in children at the Amazonas countryside.

Ingestion of small objects by children became worrisome as the number of cases increased over the years. Consequences such as intestinal obstruction or even infection of the gastrointestinal tract are becoming more frequent since even in majority cases there is having only an expectant conduct, complications can occur. Also, the combination of two elements such as punctiform metallic materials and imams cause fistula formation in loops with greater ease, granulomas in the intestines, perforation, and can progress to peritonitis or abscesses, as well as having considerable morbidity.

Target-specific forebrain projections and appropriate synaptic inputs of hESC-derived dopamine neurons grafted to the midbrain of parkinsonian rats.

Dopamine (DA) neurons derived from human embryonic stem cells (hESCs) are a promising unlimited source of cells for cell replacement therapy in Parkinson's disease (PD). A number of studies have demonstrated functionality of DA neurons originating from hESCs when grafted to the striatum of rodent and non-human primate models of PD. However, several questions remain in regard to their axonal outgrowth potential and capacity to integrate into host circuitry. Here, ventral midbrain (VM) patterned hESC-derived progenitors were grafted into the midbrain of 6-hydroxydopamine-lesioned rats, and analyzed at 6, 18, and 24 weeks for a time-course evaluation of specificity and extent of graft-derived fiber outgrowth as well as potential for functional recovery. To investigate synaptic integration of the transplanted cells, we used rabies-based monosynaptic tracing to reveal the origin and extent of host presynaptic inputs to grafts at 6 weeks. The results reveal the capacity of grafted neurons to extend axonal projections toward appropriate forebrain target structures progressively over 24 weeks. The timing and extent of graft-derived dopaminergic fibers innervating the dorsolateral striatum matched reduction in amphetamine-induced rotational asymmetry in the animals where recovery could be observed. Monosynaptic tracing demonstrated that grafted cells integrate with host circuitry 6 weeks after transplantation, in a manner that is comparable with endogenous midbrain connectivity. Thus, we demonstrate that VM patterned hESC-derived progenitors grafted to midbrain have the capacity to extensively innervate appropriate forebrain targets, integrate into the host circuitry and that functional recovery can be achieved when grafting fetal or hESC-derived DA neurons to the midbrain.

IAP-Based Cell Sorting Results in Homogeneous Transplantable Dopaminergic Precursor Cells Derived from Human Pluripotent Stem Cells.

Human pluripotent stem cell (hPSC)-derived mesencephalic dopaminergic (mesDA) neurons can relieve motor deficits in animal models of Parkinson's disease (PD). Clinical translation of differentiation protocols requires standardization of production procedures, and surface-marker-based cell sorting is considered instrumental for reproducible generation of defined cell products. Here, we demonstrate that integrin-associated protein (IAP) is a cell surface marker suitable for enrichment of hPSC-derived mesDA progenitor cells. Immunomagnetically sorted IAP+ mesDA progenitors showed increased expression of ventral midbrain floor plate markers, lacked expression of pluripotency markers, and differentiated into mature dopaminergic (DA) neurons in vitro. Intrastriatal transplantation of IAP+ cells sorted at day 16 of differentiation in a rat model of PD resulted in functional recovery. Grafts from sorted IAP+ mesDA progenitors were more homogeneous in size and DA neuron density. Thus, we suggest IAP-based sorting for reproducible prospective enrichment of mesDA progenitor cells in clinical cell replacement strategies.

Predictive Markers Guide Differentiation to Improve Graft Outcome in Clinical Translation of hESC-Based Therapy for Parkinson's Disease.

Stem cell treatments for neurodegenerative diseases are expected to reach clinical trials soon. Most of the approaches currently under development involve transplantation of immature progenitors that subsequently undergo phenotypic and functional maturation in vivo, and predicting the long-term graft outcome already at the progenitor stage remains a challenge. Here, we took an unbiased approach to identify predictive markers expressed in dopamine neuron progenitors that correlate with graft outcome in an animal model of Parkinson's disease through gene expression analysis of >30 batches of grafted human embryonic stem cell (hESC)-derived progenitors. We found that many of the commonly used markers did not accurately predict in vivo subtype-specific maturation. Instead, we identified a specific set of markers associated with the caudal midbrain that correlate with high dopaminergic yield after transplantation in vivo. Using these markers, we developed a good manufacturing practice (GMP) differentiation protocol for highly efficient and reproducible production of transplantable dopamine progenitors from hESCs.