Highly scalable and standardized organ-on-chip platform with TEER for biological barrier modeling.

The development of new therapies is hampered by the lack of predictive, and patient-relevant in vitro models. Organ-on-chip (OOC) technologies can potentially recreate physiological features and hold great promise for tissue and disease modeling. However, the non-standardized design of these chips and perfusion control systems has been a barrier to quantitative high-throughput screening (HTS). Here we present a scalable OOC microfluidic platform for applied kinetic in vitro assays (AKITA) that is applicable for high, medium, and low throughput. Its standard 96-well plate and 384-well plate layouts ensure compatibility with existing laboratory workflows and high-throughput data collection and analysis tools. The AKITA plate is optimized for the modeling of vascularized biological barriers, primarily the blood-brain barrier, skin, and lung, with precise flow control on a custom rocker. The integration of trans-epithelial electrical resistance (TEER) sensors allows rapid and repeated monitoring of barrier integrity over long time periods. Together with automated liquid handling and compound permeability testing analyses, we demonstrate the flexibility of the AKITA platform for establishing human-relevant models for preclinical drug and precision medicine's efficacy, toxicity, and permeability under near-physiological conditions.

Duration of gestation and mode of delivery affect the genes of transepithelial sodium transport in pulmonary adaptation.

BACKGROUND: Respiratory distress due to inadequate lung liquid clearance is a significant problem in infants delivered late preterm or early term, especially by elective cesarean delivery (CD). Lung liquid clearance depends on epithelial ion transport and in animals is induced by glucocorticoids. OBJECTIVES: In newborn late preterm and term infants to study airway epithelial gene expressions of epithelial sodium channel (ENaC), and the serum and glucocorticoid-inducible kinase 1 (SGK1), and their association with cortisol, mode of delivery, and gestational age (GA). METHODS: Infants were delivered at 35(0/7)-41(6/7) weeks. Cortisol in umbilical cord plasma was analyzed with liquid chromatography-tandem mass spectrometry. ENaC and SGK1 mRNAs in airway epithelial cells obtained within 3 h and at 1 day postnatally were quantified with real-time PCR. RESULTS: ENaC and SGK1 mRNAs were significantly lower in late preterm and early term infants than in those ≥ 39(0/7) weeks. Significant correlations existed between both ENaC and SGK1 and cord cortisol and GA. In term infants, SGK1 mRNA at 1.5 h was higher after vaginal delivery than elective CD. CONCLUSIONS: In late preterm and early term infants, low expression of ENaC and SGK1 may parallel insufficient lung liquid clearance predisposing to respiratory distress. Lower SGK1 expression after term CD could translate into insufficient sodium and lung liquid absorption. The findings demonstrate a central role for cortisol in regulation of ENaC and potentially perinatal sodium and lung liquid clearance.

Antenatal betamethasone associates with transient immunodepression in very low birth weight infants.

BACKGROUND: Antenatal betamethasone (BM) treatment for mothers at risk for premature delivery is effective in reducing neonatal morbidity. Immunodepression, defined as monocyte human leukocyte antigen (HLA)-DR expression <60%, is common in patients in intensive care. In very low birth weight (VLBW) infants, immunodepression correlates with gestational age and may predispose to infections. OBJECTIVES: To evaluate whether timing of antenatal BM associates with immunodepression in VLBW infants. METHODS: We determined monocyte HLA-DR expression by flow cytometry and measured 13 cytokines, cortisol, and BM in plasma from 56 VLBW infants. We calculated total glucocorticoid index as the sum of BM and cortisol at the ratio 33.3:1. RESULTS: HLA-DR expression both in cord (R(2) = 0.175, p = 0.033, n = 26) and on day 1 (R(2) = 0.125, p = 0.011, n = 51) showed an association with timing of BM. A short interval from BM to birth induced more pronounced and prolonged immunodepression, with lower HLA-DR% on postnatal day 7. On day 3, 25 infants (45%) met the criteria of immunodepression. HLA-DR expression correlated negatively with total glucocorticoid index (cord: R(2) = -0.573, p = 0.003, n = 13; day 1: R(2) = -0.213, p = 0.008, n = 32). Elapsed time from maternal BM correlated positively with concentrations of cytokines IL-6 and IL-10 on day 1. CONCLUSIONS: In VLBW infants, antenatal BM associated with transient immunodepression in a time-dependent manner. Suppression of both anti- and proinflammatory cytokines occurred. These effects may lead to an increased risk for later infections.

Coordinated release of tissue factor and tissue factor pathway inhibitor in VLBW infants.

AIM: Tissue factor (TF), a mediator between coagulation and inflammation, is upregulated in alveolar compartment and circulation in very low birthweight (VLBW) infants. We investigated the contribution of TF to systemic regulation of coagulation in VLBW infants. METHODS: We measured TF, total and free tissue factor pathway inhibitor (TFPIt, TFPIf), prothrombin fragment (F1 + 2), and thrombin-antithrombin complexes (TAT) in plasma from 51 VLBW infants during their first week of life. RESULTS: F1 + 2 in cord plasma was high (1385 pmol/mL) and decreased postnatally to 17% (p = 0.002). TAT decreased from a high cord concentration to 3% postnatally (p < 0.001). Plasma TF increased and peaked on day 3, showing no correlation with F1 + 2 or TAT. TFPIt and TFPIf increased postnatally, correlating with TF (day 1 TFPIf: R = 0.595, p < 0.001, day 3 TFPIf: R = 0.582, p < 0.001). Based on the TF/TFPIf ratio, a relative excess of plasma TF over TFPIf probably prevailed on day 3. CONCLUSIONS: In VLBW infants plasma TF fails to associate with thrombin formation. This is partly explained by release of TFPI. Despite TFPI, the newborn VLBW infant is subjected to a substantial circulating pool of TF with potential proinflammatory effects.

Low monocyte HLA-DR expression as an indicator of immunodepression in very low birth weight infants.

BACKGROUND: As a protective response to an inflammatory stimulus, the antigen-presenting molecules (human leukocyte antigen-DR (HLA-DR)) on monocytes are downregulated. If severe, the response may lead to immunodepression or immunoparalysis, associated with an increased rate of morbidity and mortality in adults. In very low birth weight (VLBW) infants, birth and intensive care present major immunological challenges. METHODS: We measured monocyte HLA-DR expression by flow cytometry and determined 13 plasma cytokines in 56 VLBW infants (gestational age (GA): 23.7-31.8 wk) and 25 controls (GA: 34.1-41.4 wk). RESULTS: HLA-DR expression decreased postnatally both in VLBW and in control infants. In VLBW infants, GA and respiratory distress syndrome (RDS) both showed associations with HLA-DR nadir on day 3, when 45% of them met the criteria of immunodepression. HLA-DR expression was lower in those infants subsequently developing infection (74 vs. 49% (day 3) and 85 vs. 68% (day 7); both P = 0.002). Interleukin (IL)-6 on day 1 was a predictor of the HLA-DR nadir. CONCLUSION: VLBW infants are in a state of immunodepression postnatally. This immunodepression correlated with GA and was a predisposing factor for late infections. The downregulation of HLA-DR during RDS probably indicates an RDS-induced antigen load on the immune system.

High tissue factor in lungs and plasma associates with respiratory morbidity in preterm infants.

AIM: In preterm infants, inflammation and intra-alveolar fibrin formation characterize respiratory distress syndrome (RDS). Tissue factor (TF) is a link between inflammation and coagulation pathways. We investigated the relationship between TF and cytokines in preterm infants to gain information of the role of TF in the inflammatory response. METHODS: We measured TF in plasma and in tracheal aspirates and analysed TF on monocytes by flow cytometry and 13 cytokines from plasma, in 56 preterm infants (birthweight 600-1500 g) during their first week. RESULTS: Plasma TF increased and peaked on day 3 and correlated with both RDS and inversely with paO2/FIO2. On day 1, TF in tracheal aspirates was 10-fold higher than in plasma and correlated with plasma TF (4888 vs. 506 pg/mL, R = 0.692, p = 0.013, n = 12). Of main pro-inflammatory cytokines, plasma TF correlated post-natally with IL-8 and IL-6 but not with IL-1 or TNF-α. CONCLUSIONS: Respiratory morbidity associates with high TF in lungs and plasma. In sick newborn infants, upregulation of TF may be mediated by IL-6 and IL-8. High TF and pro-inflammatory cytokines may together participate in the pathogenesis of pulmonary and extrapulmonary injury in preterm infants through pro-inflammatory mechanisms.