Neonatology/Paediatrics – Guidelines on Parenteral Nutrition, Chapter 13

There are special challenges in implementing parenteral nutrition (PN) in paediatric patients, which arises from the wide range of patients, ranging from extremely premature infants up to teenagers weighing up to and over 100 kg, and their varying substrate requirements. Age and maturity-related changes of the metabolism and fluid and nutrient requirements must be taken into consideration along with the clinical situation during which PN is applied. The indication, the procedure as well as the intake of fluid and substrates are very different to that known in PN-practice in adult patients, e.g. the fluid, nutrient and energy needs of premature infants and newborns per kg body weight are markedly higher than of older paediatric and adult patients. Premature infants <35 weeks of pregnancy and most sick term infants usually require full or partial PN. In neonates the actual amount of PN administered must be calculated (not estimated). Enteral nutrition should be gradually introduced and should replace PN as quickly as possible in order to minimise any side-effects from exposure to PN. Inadequate substrate intake in early infancy can cause long-term detrimental effects in terms of metabolic programming of the risk of illness in later life. If energy and nutrient demands in children and adolescents cannot be met through enteral nutrition, partial or total PN should be considered within 7 days or less depending on the nutritional state and clinical conditions

CD20 and CD19 targeted vectors induce minimal activation of resting B lymphocytes

B lymphocytes are an important cell population of the immune system. However, until recently it was not possible to transduce resting B lymphocytes with retro- or lentiviral vectors, making them unsusceptible for genetic manipulations by these vectors. Lately, we demonstrated that lentiviral vectors pseudotyped with modified measles virus (MV) glycoproteins hemagglutinin, responsible for receptor recognition, and fusion protein were able to overcome this transduction block. They use either the natural MV receptors, CD46 and signaling lymphocyte activation molecule (SLAM), for cell entry (MV-LV) or the vector particles were further modified to selectively enter via the CD20 molecule, which is exclusively expressed on B lymphocytes (CD20-LV). It has been shown previously that transduction by MV-LV does not induce B lymphocyte activation. However, if this is also true for CD20-LV is still unknown. Here, we generated a vector specific for another B lymphocyte marker, CD19, and compared its ability to transduce resting B lymphocytes with CD20-LV. The vector (CD19ds-LV) was able to stably transduce unstimulated B lymphocytes, albeit with a reduced efficiency of about 10% compared to CD20-LV, which transduced about 30% of the cells. Since CD20 as well as CD19 are closely linked to the B lymphocyte activation pathway, we investigated if engagement of CD20 or CD19 molecules by the vector particles induces activating stimuli in resting B lymphocytes. Although, activation of B lymphocytes often involves calcium influx, we did not detect elevated calcium levels. However, the activation marker CD71 was substantially up-regulated upon CD20-LV transduction and most importantly, B lymphocytes transduced with CD20-LV or CD19ds-LV entered the G1b phase of cell cycle, whereas untransduced or MV-LV transduced B lymphocytes remained in G0. Hence, CD20 and CD19 targeting vectors induce activating stimuli in resting B lymphocytes, which most likely renders them susceptible for lentiviral vector transduction

How Do Hospital Pharmacists Approach Substitution of Nanomedicines? Insights from a Qualitative Pilot Study and a Quantitative Market Research Analysis in Five European Countries

We conducted research to assess hospital pharmacists’ familiarity with/interpretation of data requirements for the different regulatory approval frameworks and the impact of this on their approach to substitution in the formulary. The online questionnaire included a small molecule (acetylsalicylic acid—follow-ons approved via the generic pathway), two biologic drugs (insulin glargine and etanercept—follow-ons approved via the biosimilar pathway), a non-biologic complex drug (NBCD; glatiramer acetate—follow-ons approved via the hybrid pathway) and a nanomedicine, ferric carboxymaltose (no follow-ons approved as yet). The study was conducted in two phases: an initial qualitative pilot study with 30 participants, followed by a quantitative stage involving 201 pharmacists from five European countries. Most expected negligible safety/efficacy differences between reference and follow-on products. Head-to-head clinical data showing therapeutic equivalence as a prerequisite for reference product/follow-on substitution was perceived to be needed most for biologics (47%), followed by NBCDs (44%)/nanomedicines (39%) and small molecules (23%). Overall, 28% did not know the data requirements for follow-on approval via the hybrid pathway; 16% were familiar with this pathway, compared with 50% and 55% for the generic and biosimilar pathways, respectively. Overall, 19% of respondents thought the European Medicines Agency (EMA) was responsible for defining the substitutability of follow-ons. Education is required to increase hospital pharmacist’s knowledge of regulatory approval frameworks and their relevance to substitution practices

Restriction of HIV-1 Replication in Monocytes Is Abolished by Vpx of SIVsmmPBj

Background: Human primary monocytes are refractory to infection with the human immunodeficiency virus 1 (HIV-1) or transduction with HIV-1-derived vectors. In contrast, efficient single round transduction of monocytes is mediated by vectors derived from simian immunodeficiency virus of sooty mangabeys (SIVsmmPBj), depending on the presence of the viral accessory protein Vpx. Methods and Findings: Here we analyzed whether Vpx of SIVsmmPBj is sufficient for transduction of primary monocytes by HIV-1-derived vectors. To enable incorporation of PBj Vpx into HIV-1 vector particles, a HA-Vpr/Vpx fusion protein was generated. Supplementation of HIV-1 vector particles with this fusion protein was not sufficient to facilitate transduction of human monocytes. However, monocyte transduction with HIV-1-derived vectors was significantly enhanced after delivery of Vpx proteins by virus-like particles (VLPs) derived from SIVsmmPBj. Moreover, pre-incubation with Vpx-containing VLPs restored replication capacity of infectious HIV-1 in human monocytes. In monocytes of non-human primates, single-round transduction with HIV-1 vectors was enabled. Conclusion: Vpx enhances transduction of primary human and even non-human monocytes with HIV-1-derived vectors, only if delivered in the background of SIVsmmPBj-derived virus-like particles. Thus, for accurate Vpx function the presence of SIVsmmPBj capsid proteins might be required. Vpx is essential to overcome a block of early infection steps in primary monocytes

Chemical stability study of vitamins thiamine, riboflavin, pyridoxine and ascorbic acid in parenteral nutrition for neonatal use

<p>Abstract</p> <p>Background</p> <p>The objective of this work was to study the vitamins B₁, B₂, B₆and C stability in a pediatric formulation containing high amounts of calcium in the presence of organic phosphate, amino acids, glucose, sodium chloride, magnesium sulfate, pediatric vitamins and trace elements under different conditions using developed and validated analytical methods.</p> <p>Methods</p> <p>The study was carried out during 72 h with formulations packaged in recommended storage temperature (4°C) and 25°C, with and without photoprotection.</p> <p>Results</p> <p>The results showed that the methodologies used for assessing the chemical stability of vitamins B₁, B₂, B₆and C in the formulation were selective, linear, precise and accurate. The vitamins could be considered stable in the formulation during the three days of study if stored at 4°C. When stored at 25°C vitamin C presented instability after 48 h.</p> <p>Conclusion</p> <p>The pediatric formulation containing high amount of calcium in the presence of organic phosphate, amino acids, glucose, sodium chloride, magnesium sulphate, pediatric vitamins and trace elements packaged in bag-type trilaminate presented a shelf life of the 72 h, when maintained under refrigeration, between 2°C and 8°C. This shelf life was measured considering the vitamins studied. Further studies are needed including all the vitamins present in this formulation.</p

Vaccine-associated enhanced respiratory pathology in COVID-19 hamsters after T(H)2-biased immunization

Vaccine-associated enhanced respiratory disease (VAERD) is a severe complication for some respiratory infections. To investigate the potential for VAERD induction in coronavirus disease 2019 (COVID-19), we evaluate two vaccine leads utilizing a severe hamster infection model: a T helper type 1 (T(H)1)-biased measles vaccine-derived candidate and a T(H)2-biased alum-adjuvanted, non-stabilized spike protein. The measles virus (MeV)-derived vaccine protects the animals, but the protein lead induces VAERD, which can be alleviated by dexamethasone treatment. Bulk transcriptomic analysis reveals that our protein vaccine prepares enhanced host gene dysregulation in the lung, exclusively up-regulating mRNAs encoding the eosinophil attractant CCL-11, T(H)2-driving interleukin (IL)-19, or T(H)2 cytokines IL-4, IL-5, and IL-13. Single-cell RNA sequencing (scRNA-seq) identifies lung macrophages or lymphoid cells as sources, respectively. Our findings imply that VAERD is caused by the concerted action of hyperstimulated macrophages and T(H)2 cytokine-secreting lymphoid cells and potentially links VAERD to antibody-dependent enhancement (ADE). In summary, we identify the cytokine drivers and cellular contributors that mediate VAERD after T(H)2-biased vaccination

Live-attenuated vaccine sCPD9 elicits superior mucosal and systemic immunity to SARS-CoV-2 variants in hamsters

Vaccines play a critical role in combating the COVID-19 pandemic. Future control of the pandemic requires improved vaccines with high efficacy against newly emerging SARS-CoV-2 variants and the ability to reduce virus transmission. Here we compare immune responses and preclinical efficacy of the mRNA vaccine BNT162b2, the adenovirus-vectored spike vaccine Ad2-spike and the live-attenuated virus vaccine candidate sCPD9 in Syrian hamsters, using both homogeneous and heterologous vaccination regimens. Comparative vaccine efficacy was assessed by employing readouts from virus titrations to single-cell RNA sequencing. Our results show that sCPD9 vaccination elicited the most robust immunity, including rapid viral clearance, reduced tissue damage, fast differentiation of pre-plasmablasts, strong systemic and mucosal humoral responses, and rapid recall of memory T cells from lung tissue after challenge with heterologous SARS-CoV-2. Overall, our results demonstrate that live-attenuated vaccines offer advantages over currently available COVID-19 vaccines

TMPRSS2-mediated SARS-CoV-2 uptake boosts innate immune activation, enhances cytopathology, and drives convergent virus evolution.

The accessory protease transmembrane protease serine 2 (TMPRSS2) enhances severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) uptake into ACE2-expressing cells, although how increased entry impacts downstream viral and host processes remains unclear. To investigate this in more detail, we performed infection assays in engineered cells promoting ACE2-mediated entry with and without TMPRSS2 coexpression. Electron microscopy and inhibitor experiments indicated TMPRSS2-mediated cell entry was associated with increased virion internalization into endosomes, and partially dependent upon clathrin-mediated endocytosis. TMPRSS2 increased panvariant uptake efficiency and enhanced early rates of virus replication, transcription, and secretion, with variant-specific profiles observed. On the host side, transcriptional profiling confirmed the magnitude of infection-induced antiviral and proinflammatory responses were linked to uptake efficiency, with TMPRSS2-assisted entry boosting early antiviral responses. In addition, TMPRSS2-enhanced infections increased rates of cytopathology, apoptosis, and necrosis and modulated virus secretion kinetics in a variant-specific manner. On the virus side, convergent signatures of cell-uptake-dependent innate immune induction were recorded in viral genomes, manifesting as switches in dominant coupled Nsp3 residues whose frequencies were correlated to the magnitude of the cellular response to infection. Experimentally, we demonstrated that selected Nsp3 mutations conferred enhanced interferon antagonism. More broadly, we show that TMPRSS2 orthologues from evolutionarily diverse mammals facilitate panvariant enhancement of cell uptake. In summary, our study uncovers previously unreported associations, linking cell entry efficiency to innate immune activation kinetics, cell death rates, virus secretion dynamics, and convergent selection of viral mutations. These data expand our understanding of TMPRSS2's role in the SARS-CoV-2 life cycle and confirm its broader significance in zoonotic reservoirs and animal models