Juan Carlos Ulloa

Postbiotics produced by probiotic bacteria are gaining attention as multifunctional, food-derived agents with potential applications in human and animal health. This study investigates the production and biological activity of protein-rich postbiotics from Bifidobacterium adolescentis, cultivated under controlled conditions in a 3-liter bioreactor as a laboratory-scale model for functional ingredient development. Culture parameters were improved, and one representative batch was selected for biological evaluation. The postbiotic preparation was tested for cytotoxicity using MA104 (renal) and C2BBe1 (intestinal) epithelial cell lines through viability and cell death assays, confirming its safety across a range of concentrations. To assess its functional activity, we evaluated its ability to reduce rotavirus infection and preserve epithelial integrity. The postbiotic significantly reduced viral infectivity and maintained cytoskeletal architecture in infected intestinal cells, supporting its potential protective role. These findings suggest that B. adolescentis-derived postbiotics may serve as safe and biologically active compounds with potential applications in intestinal health and viral infection management.

Articular cartilage regeneration remains a major challenge due to its limited self-repair capacity. Bone marrow-derived skeletal stem cells (SSCs) and mesenchymal stem cells (MSCs) are promising candidates for cartilage engineering, although they differ in their chondrogenic potential. This study explored whether co-culturing SSCs and MSCs in three-dimensional (3D) organoid systems under cartilage physioxia (5% O₂) and chondrogenic induction could improve cartilage tissue formation. SSCs, MSCs, and SSC–MSC co-cultures were characterized for morphology, phenotype, and differentiation capacity. Organoids were generated and cultured for 10 days, followed by analysis of morphology, viability, gene expression (SOX9, RUNX2, ACAN, COL2A1, COL10A1, PRG4, and PDPN), chondrocyte-associated antigens (CD44, CD105, CD146, and PDPN), and cartilage ECM proteins (aggrecan, collagen types I, II, and X, and PRG4). SSCs showed robust chondrogenic and osteogenic potential, while MSCs exhibited a balanced multipotency. Co-culture-derived organoids enhanced chondrogenesis and reduced adipogenesis, with higher expression of cartilage-specific ECM and lower hypertrophic marker levels. These findings highlight the functional synergy between SSCs and MSCs in co-culture, promoting the formation of stable, cartilage-like structures under physioxia. The approach offers a promising strategy for generating preclinical models and advancing regenerative therapies for hyaline cartilage repair.

Articular cartilage regeneration remains a major challenge due to its limited self-repair capacity. Bone marrow-derived skeletal stem cells (SSCs) and mesenchymal stem cells (MSCs) are promising candidates for cartilage engineering, though they differ in their chondrogenic potential. This study explored whether co-culturing SSCs and MSCs in three-dimensional (3D) organoid systems under cartilage physioxia (5% O₂) and chon-drogenic induction could improve cartilage tissue formation. SSCs, MSCs, and SSC–MSC co-cultures were characterized for morphology, phenotype, and differentiation capacity. Organoids were generated and cultured for 10 days, followed by analysis of morphology, viability, gene expression (SOX9, RUNX2, ACAN, COL2A1, COL10A1, PRG4, and PDPN), chondrocyte-associated antigens (CD44, CD105, CD146, and PDPN), and cartilage ECM proteins (aggrecan, collagen types I, II, and X, and PRG4). SSCs showed robust chondrogenic and osteogenic potential, while MSCs exhibited a balanced multipotency. Co-culture-derived organoids enhanced chondrogenesis and reduced adipogenesis, with higher expression of cartilage-specific ECM and lower hy-pertrophic marker levels. These findings highlight the functional synergy between SSCs and MSCs in co-culture, promoting the formation of stable, cartilage-like struc-tures under physioxia. The approach offers a promising strategy for generating pre-clinical models and advancing regenerative therapies for hyaline cartilage repair.

Acute diarrheal disease (ADD) caused by rotavirus (RV) contributes significantly to morbidity and mortality in children under five years of age. Currently, there are no specific drugs for the treatment of RV infections. Previously, we reported the anti-rotaviral activity of the protein metabolites derived from Bifidobacterium adolescentis. In this study, our aim was to assess the impact of B. adolescentis-secreted proteins (BaSP), with anti-rotaviral activity on the human intestinal C2BBe1 cell line. We initiated the production of BaSP and subsequently confirmed its anti-rotaviral activity by counting the infectious foci using immunocytochemistry. We then exposed the C2BBe1 cells to various concentrations of BaSP (≤250 µg/mL) for 72 h. Cell viability was assessed using the MTT assay, cell monolayer integrity was monitored through transepithelial electrical resistance (TEER), and cytoskeleton architecture and tight junctions (TJs) were examined using confocal microscopy with F-actin and occludin staining. Finally, we utilized a commercial kit to detect markers of apoptosis and necrosis after 24 h of treatment. The results demonstrated that BaSP does not have adverse effects on C2BBe1 cells. These findings confirm that BaSP inhibits rotavirus infectivity and has the potential to strengthen intestinal defense against viral and bacterial infections via the paracellular route.

Viruses represent the primary etiologic agents (70–80%) of acute diarrheal disease (ADD), and rotavirus (RV) is the most relevant one. Currently, four rotavirus vaccines are available. However, these vaccines do not protect against emerging viral strains or are not available in low-income countries. To date, there are no approved drugs available against rotavirus infection. In this study, we evaluated the in vitro anti-rotaviral activity and intestinal toxicity of a phytotherapeutic prototype obtained from Achyrocline bogotensis (Kunth) DC. (PPAb); medicinal plant that contains compounds that inhibit the rotavirus replication cycle. Virucidal and viral yield reduction effects exerted by the PPAb were evaluated by immunocytochemistry and flow cytometry. Furthermore, the toxic impact of the PPAb was evaluated in polarized human intestinal epithelial C2BBe1 cells in terms of cytotoxicity, loss of cytoplasmic membrane asymmetry, and DNA fragmentation by MTT and fluorometry. PPAb concentrations under 0.49 mg/mL exerted significant virucidal and viral yield reduction activities, and concentrations under 16 mg/mL neither reduced cell viability, produced DNA fragmentation, nor compromised the C2BBe1cell membrane stability after 24-h incubation. Based on these results, the evaluated phytotherapeutic prototype of Achyrocline bogotensis might be considered as a promising alternative to treat ADD caused by rotavirus.

Claudia Camila Mejía-Cruz, Emilia Barreto-Durán, María Alejandra Pardo-Pérez, María Camila Jimenez, Julieth Rincón, Karen Vanegas, Jorge Luis Rodríguez, Luis Fernando Jaramillo-Garcia, Juan Carlos Ulloa, Rodolfo Martínez Díaz, Efrain Leal-García, Rafael Pérez-Núñez, Alfonso Barreto, Viviana M Rodríguez-Pardo. IJSC 1999;0:. https://doi.org/10.15283/ijsc18061

Achyrocline bogotensis has been traditionally used to treat infections of skin, respiratory, tract urinary and other infections, but not to treat viral gastrointestinal disease. In this study, this Colombian native medicinal plant was investigated by its in vitro anti-rotavirus and anti-astrovirus activity. Several extracts and fractions phytochemically obtained from A. bogotensis were evaluated initially for their cell toxicity on MA104 and Caco2 cells and then for their anti-rotavirus (RRV) and anti-astrovirus (Yuc8) activity following three strategies: pre-treatment of cells (blocking effect), direct viral activity (virucidal effect) and post-treatment of infected cells (reduction of viral yield post-infection). In addition qualitative chemical studies were developed for the active compounds. Non-toxic concentrations of a fraction obtained exhibited antiviral activity against both viruses characterized by a virucidal effect and by the reduction of the infectious particles produced post-infection. Steroids, sterols, terpenes, phenols, flavonoids and sesquiterpenlactones were identified qualitatively in the active fraction. A. bogotensis contains substances with in vitro antiviral activity against rotavirus and astrovirus. This study confirms their anti-microbial properties and describes by the first time its antiviral activity in vitro.

Los tejidos embebidos en parafina son muestras que reposan en los bancos de patología y almacenan importante información genética, útil para estudios retrospectivos de epidemiología y de patología molecular y para el diagnóstico de enfermedades en humanos y en animales. Dentro de las metodologías descritas para extraer ADN de tejidos en parafina, pocas resultan exitosas a la hora de obtener un material genético, con la cual se pueda amplificar fragmentos superiores a 200pb, por la técnica de PCR. Este trabajo expone la experiencia del laboratorio de biología molecular de la U.D.C.A, relacionada con la aplicación de una metodología poco utilizada para extraer ADN de tejido embebido en parafina y óptimo para amplificar por PCR un segmento de 600pb. Fueron utilizadas dos técnicas ampliamente referenciadas para este tipo de muestras, como son el protocolo de extracción fenólica y el del estuche comercial de QIAGEN^® y una tercera metodología comúnmente empleada para extraer ADN de muestras de sangre, pero no para para tejidos en parafina, como lo es la técnica de extracción por sales o salting out. La capacidad amplificable del ADN extraído, se evidenció al obtener un fragmento de 600pb de un gen constitutivo humano. Los resultados mostraron que con la extracción fenólica y el estuche comercial de QIAGEN^®, se consiguió el 21% de amplificaciones y con el método de extracción por sales, el 91%. Este trabajo muestra que una técnica no convencional de extracción de ADN a partir de tejido embebido en parafina puede ofrecer mejores resultados que metodologías tradicionales.

ResumenObjetivoDeterminar la presencia de rotavirus y norovirus en agua para consumo humano en una localidad de Bogotá.Materiales y métodosSe recolectaron ocho muestras de agua por semana en ocho lugares diferentes del predio en estudio. Para determinar la presencia del genoma viral, las muestras se ultrafiltraron y se hizo detección viral por medio de la técnica RT-PCR, amplificando un segmento del gen VP6 del rotavirus y un segmento del ORF2 del norovirus. Este estudio se llevó a cabo en una institución universitaria ubicada en una zona periurbana cercana a Bogotá, a 45km del acueducto que surte de agua al predio. La zona se encuentra rodeada por fincas con ganado lechero, centros comerciales y cuatro cementerios. El agua se recolectó en recipientes de polipropileno. En total, fueron 64 muestras de cinco litros de agua cada una.ResultadosLa presencia viral fue de 12,5%, encontrándose rotavirus en cuatro y norovirus en cuatro de las 64 muestras colectadas. Además de presentar la importancia de la presencia viral en agua para consumo humano, en este documento se discute el significado que tiene el encontrar solo segmentos virales y no la partícula viral completa e infecciosa.AbstractObjectiveTo determine the presence of rotavirus and norovirus in drinking water in a northern neighborhood in Bogotá, Colombia.Materials and methodsEight weekly samples of water were collected and ultrafiltered in order to detect the viral genome presence. The technique used for detection of genome segments was RT-PCR. VP6 gene from rotavirus and ORF2 segment from norovirus were selected in order to find the virus in the water. This study was performed in a higher education institution located in a periurban zone near Bogotá, 45km away from the aqueduct that services the area. The zone is surrounded by dairy cattle farms, malls and four cemeteries. Water was collected in clean polypropylene containers. There were a total of 64 samples of 5 liters of water each.ResultsViral segments were found in 12.5% of the samples. We found rotavirus in four samples and norovirus in another 4. Besides discussing the importance of viral contamination in drinking water, we discussed the meaning of finding only viral segments and not complete and infectious viral particles

Porcine astroviruses (PAstVs) are small RNA viruses associated with gastroenteritis. The capsid polyprotein of PAstVs, human (HAstVs) and feline (FAstVs) AstVs has a high similarity at the N-terminus before residue 415. Previous results showed the cross-detection of PAstVs and HAstVs from diarrheal samples using a commercial ELISA test that uses a monoclonal antibody to capture HAstVs, suggesting the existence of common immunoreactive epitopes between these two virus types. In this study, we seeked immunoreactive peptides located in the PAstV capsid that may be potentially used for their specific detection. The variability and hydrophobicity of a short fragment of 132 amino acids were analyzed using several capsid sequences of PAstVs and HAstVs. Peptides TATL, SLNP and IDIV were selected, synthesized and inoculated into rabbits. Pre- and hyperimmune sera were collected and their reactivity was examined by immunoassay and immunofluorescence against two wild-type strains of PAstV adapted to grow in cell culture, observing reactivity in two of the sera. Finally, the possible cross-reactivity of the sera against HAstVs was partially ruled out using HAstV8. Our data suggest that TATLGTIGSNSSGKTELEAC and IDIVVGKAATFNLKASDLSGP peptides represent immunoreactive regions useful for the specific detection of PAstVs.