Derived Time in Range and Other Metrics of Poor Glycemic Control Associated With Adverse Hospital Outcomes in Patients With Diabetes Mellitus Admitted to Non-ICU Wards at a Tertiary-Level Hospital in Colombia: A Cross-Sectional Study.

Aim: This study is aimed at assessing the prevalence of poor glycemic control using different metrics and its association with in-hospital adverse outcomes. Methods: This cross-sectional study was conducted in diabetic patients admitted to a third-level hospital in Colombia between January and July 2022. Poor glycemic control was determined using capillary glucose metrics, including mean glucose values outside the target range, derived time in range (dTIR) (100-180 mg/dL) < 70%, coefficient of variation (CV > 36%), and hypoglycemia (<70 mg/dL). Multiple regression models were adjusted for hospital outcomes based on glycemic control, as well as other sociodemographic and clinical covariates. Results: A total of 330 Hispanic patients were included. A total of 27.6% had mean glucose measurements outside the target range, 33% had a high CV, 64.8% had low dTIR, and 28.8% experienced hypoglycemia. The in-hospital mortality rate was 8.8%. An admission HbA1c level greater than 7% was linked to an increased mortality risk (p = 0.016), as well as a higher average of glucometer readings (186 mg/dL vs. 143 mg/dL; p < 0.001). A lower average of dTIR (41.0% vs. 60.0%; p < 0.001) was also associated with a higher mortality risk. Glycemic variability was correlated with an increased risk of mortality, hypoglycemia, delirium, and length of hospital stay (LOS). Conclusion: A significant number of hospitalized diabetic patients exhibit poor glycemic control, which has been found to be associated with adverse outcomes, including increased mortality. Metrics like dTIR and glycemic variability should be considered as targets for glycemic control, highlighting the need for enhanced management strategies.

A tridimensional atlas of the developing human head.

The evolution and development of the head have long captivated researchers due to the crucial role of the head as the gateway for sensory stimuli and the intricate structural complexity of the head. Although significant progress has been made in understanding head development in various vertebrate species, our knowledge of early human head ontogeny remains limited. Here, we used advanced whole-mount immunostaining and 3D imaging techniques to generate a comprehensive 3D cellular atlas of human head embryogenesis. We present detailed developmental series of diverse head tissues and cell types, including muscles, vasculature, cartilage, peripheral nerves, and exocrine glands. These datasets, accessible through a dedicated web interface, provide insights into human embryogenesis. We offer perspectives on the branching morphogenesis of human exocrine glands and unknown features of the development of neurovascular and skeletomuscular structures. These insights into human embryology have important implications for understanding craniofacial defects and neurological disorders and advancing diagnostic and therapeutic strategies.

In Utero Electroporation of Multiaddressable Genome-Integrating Color (MAGIC) Markers to Individualize Cortical Mouse Astrocytes.

Protoplasmic astrocytes (PrA) located in the mouse cerebral cortex are tightly juxtaposed, forming an apparently continuous three-dimensional matrix at adult stages. Thus far, no immunostaining strategy can single them out and segment their morphology in mature animals and over the course of corticogenesis. Cortical PrA originate from progenitors located in the dorsal pallium and can easily be targeted using in utero electroporation of integrative vectors. A protocol is presented here to label these cells with the multiaddressable genome-integrating color (MAGIC) Markers strategy, which relies on piggyBac/Tol2 transposition and Cre/lox recombination to stochastically express distinct fluorescent proteins (blue, cyan, yellow, and red) addressed to specific subcellular compartments. This multicolor fate mapping strategy enables to mark in situ nearby cortical progenitors with combinations of color markers prior to the start of gliogenesis and to track their descendants, including astrocytes, from embryonic to adult stages at the individual cell level. Semi-sparse labeling achieved by adjusting the concentration of electroporated vectors and color contrasts provided by the Multiaddressable Genome-Integrating Color Markers (MAGIC Markers or MM) enable to individualize astrocytes and single out their territory and complex morphology despite their dense anatomical arrangement. Presented here is a comprehensive experimental workflow including the details of the electroporation procedure, multichannel image stacks acquisition by confocal microscopy, and computer-assisted three-dimensional segmentation that will enable the experimenter to assess individual PrA volume and morphology. In summary, electroporation of MAGIC Markers provides a convenient method to individually label numerous astrocytes and gain access to their anatomical features at different developmental stages. This technique will be useful to analyze cortical astrocyte morphological properties in various mouse models without resorting to complex crosses with transgenic reporter lines.

Cortical astrocytes develop in a plastic manner at both clonal and cellular levels.

Astrocytes play essential roles in the neural tissue where they form a continuous network, while displaying important local heterogeneity. Here, we performed multiclonal lineage tracing using combinatorial genetic markers together with a new large volume color imaging approach to study astrocyte development in the mouse cortex. We show that cortical astrocyte clones intermix with their neighbors and display extensive variability in terms of spatial organization, number and subtypes of cells generated. Clones develop through 3D spatial dispersion, while at the individual level astrocytes acquire progressively their complex morphology. Furthermore, we find that the astroglial network is supplied both before and after birth by ventricular progenitors that scatter in the neocortex and can give rise to protoplasmic as well as pial astrocyte subtypes. Altogether, these data suggest a model in which astrocyte precursors colonize the neocortex perinatally in a non-ordered manner, with local environment likely determining astrocyte clonal expansion and final morphotype.

Diffusion-weighted magnetic resonance spectroscopy enables cell-specific monitoring of astrocyte reactivity in vivo.

Reactive astrocytes exhibit hypertrophic morphology and altered metabolism. Deciphering astrocytic status would be of great importance to understand their role and dysregulation in pathologies, but most analytical methods remain highly invasive or destructive. The diffusion of brain metabolites, as non-invasively measured using diffusion-weighted magnetic resonance spectroscopy (DW-MRS) in vivo, depends on the structure of their micro-environment. Here we perform advanced DW-MRS in a mouse model of reactive astrocytes to determine how cellular compartments confining metabolite diffusion are changing. This reveals myo-inositol as a specific intra-astrocytic marker whose diffusion closely reflects astrocytic morphology, enabling non-invasive detection of astrocyte hypertrophy (subsequently confirmed by confocal microscopy ex vivo). Furthermore, we measure massive variations of lactate diffusion properties, suggesting that intracellular lactate is predominantly astrocytic under control conditions, but predominantly neuronal in case of astrocyte reactivity. This indicates massive remodeling of lactate metabolism, as lactate compartmentation is tightly linked to the astrocyte-to-neuron lactate shuttle mechanism.

Efficient GPU-based Monte-Carlo simulation of diffusion in real astrocytes reconstructed from confocal microscopy.

The primary goal of this work is to develop an efficient Monte-Carlo simulation of diffusion-weighted signal in complex cellular structures, such as astrocytes, directly derived from confocal microscopy. In this study, we first use an octree structure for spatial decomposition of surface meshes. Octree structure and radius-search algorithm help to quickly identify the faces that particles can possibly encounter during the next time step, thus speeding up the Monte-Carlo simulation. Furthermore, we propose to use a three-dimensional binary marker to describe the complex cellular structure and optimize the particle trajectory simulation. Finally, a GPU-based version of these two approaches is implemented for more efficient modeling. It is shown that the GPU-based binary marker approach yields unparalleled performance, opening up new possibilities to better understand intracellular diffusion, validate diffusion models, and create dictionaries of intracellular diffusion signatures.

Can we detect the effect of spines and leaflets on the diffusion of brain intracellular metabolites?

Prior models used to clarify which aspects of tissue microstructure mostly affect intracellular diffusion and corresponding diffusion-weighted magnetic resonance (DW-MR) signal have focused on relatively simple geometrical descriptions of the cellular microenvironment (spheres, randomly oriented cylinders, etc…), neglecting finer morphological details which may have an important role. Some types of neurons present high density of spines; and astrocytes and macroglial cells processes present leaflets, which may all impact the diffusion process. Here, we use Monte-Carlo simulations of many particles diffusing in cylindrical compartments with secondary structures mimicking spines and leaflets of neuronal and glial cell fibers, to investigate to what extent the diffusion-weighted signal of intracellular molecules is sensitive to spines/leaflets density and length. In order to study the specificity of DW-MR signal to these kinds of secondary structures, beading-like geometry is simulated as "control" deviation from smooth cylinder too. Results suggest that: a) the estimated intracellular tortuosity increases as spines/leaflets density or length (beading amplitude) increase; b) the tortuosity limit is reached for diffusion time td>200 ms for metabolites and td>70 ms for water molecules, suggesting that the effects of these finer morphological details are negligible at td longer than these threshold values; c) fiber diameter is overestimated, while intracellular diffusivity is underestimated, when simple geometrical models based on hollow smooth cylinders are used; d) apparent surface-to-volume, S/V, ratio estimated by linear fit of high frequency OG data appears to be an excellent estimation of the actual S/V ratio, even in the presence of secondary structures, and it increases as spines and leaflets density or length increase (while decreasing as beadings amplitude increases). Comparison between numerical simulations and multimodal metabolites DW-MRS experiments in vivo in mouse brain shows that these fine structures may affect the DW-MRS signal and the derived diffusion metrics consistently with their expected density and geometrical features. This work suggests that finer structures of cell morphology have non-negligible effects on intracellular molecules' diffusion that may be measured by using multimodal DW-MRS approaches, stimulating future developments and applications.

Long-Term Efficacy and Safety of Sensor Augmented Insulin Pump Therapy with Low-Glucose Suspend Feature in Patients with Type 1 Diabetes.

BACKGROUND: Literature supports short-term efficacy and safety of Sensor Augmented Insulin Pump (SAP) therapy in patients with type 1 diabetes (T1D). However, no data are available showing long-term outcomes. Our study describes the long-term outcomes of SAP therapy with low-glucose suspend feature (SAP+LGS) in hypoglycemia in a Colombian population with T1D and hypoglycemia. METHODS: A cohort study was conducted with T1D patients receiving SAP+LGS therapy who initiated this therapy because of hypoglycemia at San Ignacio University Hospital diabetes center in Bogotá, Colombia. Glycated hemoglobin (A1c) was assessed at least every 6 months, severe hypoglycemia (SH) and hypoglycemia unawareness (HU) incidence yearly. Adherence to therapy was also evaluated. RESULTS: One hundred eleven patients were included in the analysis. Total daily insulin dose was reduced during follow-up (mean difference -0.22 U/kg; 95% confidence interval [CI] -0.18 to -0.26; P < 0.001). A1c levels were reduced from a baseline value of 8.8% ± 1.9% to 7.5% ± 1.0% at 5 months (mean difference -1.3%; 95% CI -1.09 to -1.50; P < 0.001) and 7.1% ± 0.8% (mean difference -1.7%; 95% CI -1.59 to -1.90; P < 0.001) at the end of follow-up (47 months on average). The incidence of SH and HU episodes decreased significantly since the first year, and this effect was maintained over time (P < 0.001). CONCLUSIONS: SAP+LGS therapy in T1D patients with hypoglycemia led to a significant and sustained decrease in A1c during long periods of follow-up, as well as a significant reduction in SH and HU. Future randomized clinical trials are desired.

Insulin Regulates Astrocytic Glucose Handling Through Cooperation With IGF-I.

Brain activity requires a flux of glucose to active regions to sustain increased metabolic demands. Insulin, the main regulator of glucose handling in the body, has been traditionally considered not to intervene in this process. However, we now report that insulin modulates brain glucose metabolism by acting on astrocytes in concert with IGF-I. The cooperation of insulin and IGF-I is needed to recover neuronal activity after hypoglycemia. Analysis of underlying mechanisms show that the combined action of IGF-I and insulin synergistically stimulates a mitogen-activated protein kinase/protein kinase D pathway resulting in translocation of GLUT1 to the cell membrane through multiple protein-protein interactions involving the scaffolding protein GAIP-interacting protein C terminus and the GTPase RAC1. Our observations identify insulin-like peptides as physiological modulators of brain glucose handling, providing further support to consider the brain as a target organ in diabetes.

The insulin-like growth factor I receptor regulates glucose transport by astrocytes.

Previous findings indicate that reducing brain insulin-like growth factor I receptor (IGF-IR) activity promotes ample neuroprotection. We now examined a possible action of IGF-IR on brain glucose transport to explain its wide protective activity, as energy availability is crucial for healthy tissue function. Using (18) FGlucose PET we found that shRNA interference of IGF-IR in mouse somatosensory cortex significantly increased glucose uptake upon sensory stimulation. In vivo microscopy using astrocyte specific staining showed that after IGF-IR shRNA injection in somatosensory cortex, astrocytes displayed greater increases in glucose uptake as compared to astrocytes in the scramble-injected side. Further, mice with the IGF-IR knock down in astrocytes showed increased glucose uptake in somatosensory cortex upon sensory stimulation. Analysis of underlying mechanisms indicated that IGF-IR interacts with glucose transporter 1 (GLUT1), the main facilitative glucose transporter in astrocytes, through a mechanism involving interactions with the scaffolding protein GIPC and the multicargo transporter LRP1 to retain GLUT1 inside the cell. These findings identify IGF-IR as a key modulator of brain glucose metabolism through its inhibitory action on astrocytic GLUT1 activity. GLIA 2016;64:1962-1971.

Conversión de tuberculina en trabajadores del Hospital Universitario San Ignacio y asociación con características demográficas y laborales / Tuberculin skin test conversion and association with occupation and demographic characteristics in workers at San Ignacio University Hospital

Objetivos: Establecer la prevalencia de positividad, la tasa de conversión de la tuberculina en trabajadores de nuestro hospital y describir las características demográficas y laborales asociadas a esto. Materiales y Métodos Estudio observacional descriptivo en una cohorte de trabajadores del hospital a los que se les realizó la prueba de tuberculina. Se definió la positividad de la prueba de tuberculina como un resultado mayor o igual a 10 mm y la conversión como un aumento de 6 o 10 mm con respecto a la prueba inicial. Resultados Se encontró una prevalencia de 23,7 % y una incidencia de conversión de 13,6 % para el punto de corte mayor de 10 mm y 23 % para el punto de corte de 6 mm. La edad de los sujetos estuvo relacionada a conversión, mientras que no se encontró relación con la ocupación. Conclusión La probabilidad de transmisión de la tuberculosis en trabajadores del hospital es mayor al de la población general. Deben ser implementadas medidas de promoción y prevención para disminuir la transmisión e incrementar el conocimiento de la tuberculosis asociada al cuidado de la salud en los trabajadores.(AU)

Objectives To establish the prevalence of positivity and conversion rate of the tuberculin skin test in workers of our hospital, and to describe the related demographic and occupational characteristics. Materials and Methods An observational, descriptive study was conducted in a cohort of hospital workers who underwent the tuberculin skin test. The positivity of the test was defined as a result of greater than or equal to 10 mm, and conversion was defined as an increase of 6 or 10 mm with respect to the initial test. Results Prevalence of 23.7 % and incidence of conversion of 13.6 % for the major cut-off point of 10 mm, and 23 % for the lesser cut-off point of 6 mm, was found. The age of the subjects was related to the conversion, but there was no relation with occupation. Conclusions The probability of tuberculosis transmission in health-care workers is higher than in the general population. Promotion and prevention measures must be implemented to decrease the transmission and to increase awareness of tuberculosis related to occupational activities.(AU)

[Tuberculin skin test conversion and association with occupation and demographic characteristics in workers at San Ignacio University Hospital]. / Conversión de tuberculina en trabajadores del Hospital Universitario San Ignacio y asociación con características demográficas y laborales.

Objectives To establish the prevalence of positivity and conversion rate of the tuberculin skin test in workers of our hospital, and to describe the related demographic and occupational characteristics. Materials and Methods An observational, descriptive study was conducted in a cohort of hospital workers who underwent the tuberculin skin test. The positivity of the test was defined as a result of greater than or equal to 10 mm, and conversion was defined as an increase of 6 or 10 mm with respect to the initial test. Results Prevalence of 23.7 % and incidence of conversion of 13.6 % for the major cut-off point of 10 mm, and 23 % for the lesser cut-off point of 6 mm, was found. The age of the subjects was related to the conversion, but there was no relation with occupation. Conclusions The probability of tuberculosis transmission in health-care workers is higher than in the general population. Promotion and prevention measures must be implemented to decrease the transmission and to increase awareness of tuberculosis related to occupational activities.

The Trypanosoma brucei gambiense secretome impairs lipopolysaccharide-induced maturation, cytokine production, and allostimulatory capacity of dendritic cells.

Trypanosoma brucei gambiense, a parasitic protozoan belonging to kinetoplastids, is the main etiological agent of human African trypanosomiasis (HAT), or sleeping sickness. One major characteristic of this disease is the dysregulation of the host immune system. The present study demonstrates that the secretome (excreted-secreted proteins) of T. b. gambiense impairs the lipopolysaccharide (LPS)-induced maturation of murine dendritic cells (DCs). The upregulation of major histocompatibility complex class II, CD40, CD80, and CD86 molecules, as well as the secretion of cytokines such as tumor necrosis factor alpha, interleukin-10 (IL-10), and IL-6, which are normally released at high levels by LPS-stimulated DCs, is significantly reduced when these cells are cultured in the presence of the T. b. gambiense secretome. Moreover, the inhibition of DC maturation results in the loss of their allostimulatory capacity, leading to a dramatic decrease in Th1/Th2 cytokine production by cocultured lymphocytes. These results provide new insights into a novel efficient immunosuppressive mechanism directly involving the alteration of DC function which might be used by T. b. gambiense to interfere with the host immune responses in HAT and promote the infectious process.

Use of OmpU porins for attachment and invasion of Crassostrea gigas immune cells by the oyster pathogen Vibrio splendidus.

OmpU porins are increasingly recognized as key determinants of pathogenic host Vibrio interactions. Although mechanisms remain incompletely understood, various species, including the human pathogen Vibrio cholera, require OmpU for host colonization and virulence. We have shown previously that OmpU is essential for virulence in the oyster pathogen Vibrio splendidus LGP32. Here, we showed that V. splendidus LGP32 invades the oyster immune cells, the hemocytes, through subversion of host-cell actin cytoskeleton. In this process, OmpU serves as an adhesin/invasin required for ß-integrin recognition and host cell invasion. Furthermore, the major protein of oyster plasma, the extracellular superoxide dismutase Cg-EcSOD, is used as an opsonin mediating the OmpU-promoted phagocytosis through its RGD sequence. Finally, the endocytosed bacteria were found to survive intracellularly, evading the host defense by preventing acidic vacuole formation and limiting reactive oxygen species production. We conclude that (i) V. splendidus is a facultative intracellular pathogen that manipulates host defense mechanisms to enter and survive in host immune cells, and (ii) that OmpU is a major determinant of host cell invasion in Vibrio species, used by V. splendidus LGP32 to attach and invade oyster hemocytes through opsonisation by the oyster plasma Cg-EcSOD.

Expression and immunogenicity of the mycobacterial Ag85B/ESAT-6 antigens produced in transgenic plants by elastin-like peptide fusion strategy.

This study explored a novel system combining plant-based production and the elastin-like peptide (ELP) fusion strategy to produce vaccinal antigens against tuberculosis. Transgenic tobacco plants expressing the mycobacterial antigens Ag85B and ESAT-6 fused to ELP (TBAg-ELP) were generated. Purified TBAg-ELP was obtained by the highly efficient, cost-effective, inverse transition cycling (ICT) method and tested in mice. Furthermore, safety and immunogenicity of the crude tobacco leaf extracts were assessed in piglets. Antibodies recognizing mycobacterial antigens were produced in mice and piglets. A T-cell immune response able to recognize the native mycobacterial antigens was detected in mice. These findings showed that the native Ag85B and ESAT-6 mycobacterial B- and T-cell epitopes were conserved in the plant-expressed TBAg-ELP. This study presents the first results of an efficient plant-expression system, relying on the elastin-like peptide fusion strategy, to produce a safe and immunogenic mycobacterial Ag85B-ESAT-6 fusion protein as a potential vaccine candidate against tuberculosis.

Sequence diversity and differential expression of Tc52 immuno-regulatory protein in Trypanosoma cruzi: potential implications in the biological variability of strains.

Trypanosoma cruzi is highly heterogeneous in terms of genetics and biological properties. To explore the diversity of T. cruzi, we focused our study on the T. cruzi Tc52 protein playing a critical immunosuppressive role during infection. Sequence variability and expression levels of this virulence factor were analysed in various strains. Among the 40 amino acid substitutions detected in the Tc52 coding sequences, three substitutions may have an impact on protein activity or function, as two are localized in sites involved in the glutathione binding and the third is present in the region bearing immunomodulatory function. This sequence variability was consistent with the genetic subdivisions of T. cruzi. Moreover, we observed that the level of Tc52 transcripts and proteins varied between the different strains, but we did not find a significant correlation between Tc52 expression and the phylogeny of the parasite. Thus, both diversity in the sequences and differences in the expression levels of Tc52 protein may be involved in the biological variability of T. cruzi, especially in virulence and immunosuppression properties of T. cruzi strains.

Trypanosoma cruzi carrying a targeted deletion of a Tc52 protein-encoding allele elicits attenuated Chagas' disease in mice.

The intracellular protozoan parasite Trypanosoma cruzi is the etiological agent of Chagas' disease. We have previously characterized a T. cruzi virulence factor named Tc52 sharing structural and functional properties with the thioredoxin and glutaredoxin protein family. Single mutant parasite clones (Tc52(+/-)) exhibiting low virulence in vitro and in vivo were obtained by targeted Tc52 gene replacement. In this report, we have extended our study to analyze the immune response and the disease phenotype in Tc52(+/-)-infected BALB/c mice, during the acute and chronic phases of the disease. Significantly lower parasitemia were found in Tc52(+/-)-infected mice, as compared to wild-type parasite (WT)-infected ones. However, the expansion of all classes of lymphocytes and macrophages was similar for both clones. Furthermore, except for IgG2b levels which were higher in the case of WT-infected mice, all classes of Ig presented no significant difference for WT and Tc52(+/-)-infected animals. Interestingly, a lack of suppression of IL-2 production and of T-cell proliferation inhibition was observed in the case of spleen cells from Tc52(+/-)-infected mice. Finally, the pattern of inflammation process was different and characterized as diffused in the case of Tc52(+/-)-infected mice, or presenting numerous foci in the case of WT-infected mice. Localization of the Tc52 protein in tissue sections and infected heart cell primary cultures by immunofluorescence and immunogold labeling, respectively, revealed the presence of Tc52 at the amastigote surface and associated to aggregates within host cell vesicles. Taken together, these results reinforce the notion of Tc52 being a virulence factor playing a role in the phenotype of the immune response associated to the infection and on the course of the disease.