Draining the Pleural Space: Lymphatic Vessels Facing the Most Challenging Task.

Lymphatic vessels exploit the mechanical stresses of their surroundings together with intrinsic rhythmic contractions to drain lymph from interstitial spaces and serosal cavities to eventually empty into the blood venous stream. This task is more difficult when the liquid to be drained has a very subatmospheric pressure, as it occurs in the pleural cavity. This peculiar space must maintain a very low fluid volume at negative hydraulic pressure in order to guarantee a proper mechanical coupling between the chest wall and lungs. To better understand the potential for liquid drainage, the key parameter to be considered is the difference in hydraulic pressure between the pleural space and the lymphatic lumen. In this review we collected old and new findings from in vivo direct measurements of hydraulic pressures in anaesthetized animals with the aim to better frame the complex physiology of diaphragmatic and intercostal lymphatics which drain liquid from the pleural cavity.

Late-term fetuses with reduced umbilical vein blood flow volume: An under-recognized population at increased risk of growth restriction.

OBJECTIVES: To investigate the umbilical vein and uterine arteries blood flow volume (UV-Q, UtA-Q) in late-term pregnancies. STUDY DESIGN: This was a prospective observational cohort study of singleton pregnancies ≥40 + 0 weeks in which UV-Q and UtA-Q, both absolute and normalized for estimated fetal weight (EFW) values, were evaluated in relation to AC drop of ≥20 percentiles from 20 weeks to term, Doppler signs of fetal cerebral blood flow redistribution and composite adverse perinatal outcome. The presence of neonatal hypoglycaemia and the need of formula milk supplementation were also examined. RESULTS: The study population comprised 200 women. Fetuses with AC drop (n = 34) had a significantly lower UV-Q and UV-Q/EFW than fetuses without AC drop (n = 166): median UV-Q 184 ml/min (IQR 143-225) vs 233 ml/min (IQR 181-277), p = 0.0006; median UV-Q/EFW 55 ml/min/kg (IQR 42-66) vs 63 ml/min/kg (IQR 48-74), p = 0.03. Fetuses with cerebral blood flow redistribution (n = 48) had a significantly lower UV-Q and UV-Q/EFW than those without (n = 134): median UV-Q 210 ml/min (IQR 155-263) vs 236 ml/min (IQR 184-278), p = 0.04; median UV-Q/EFV 58 ml/min/kg (IQR 45-70) vs 65 ml/min/kg (IQR 50-76), p = 0.04. There was a significant moderate correlation between middle cerebral artery pulsatility index (MCA-PI) and UV-Q and UV-Q/EFW (Spearman Rho -0.20 and -0.20; p = 0.008 and p = 0.006). CONCLUSIONS: The umbilical vein blood flow volume might have a potential role to identify fetuses with stunted growth in late-term pregnancies.

Evidence of bacterial DNA presence in chorionic villi and amniotic fluid in the first and second trimester of pregnancy.

The sterile-womb dogma in uncomplicated pregnancy has been lively debated. Data regarding the in utero microbiome environment are based mainly on studies performed at the time of delivery. Aim: To determine whether human placenta and amniotic fluid are populated by a bacterial microbiota in the first and second trimesters of pregnancy. Materials & methods: We analyzed by next-generation sequencing method 24 and 29 samples from chorionic villus sampling (CVS) and amniocentesis (AC), respectively. The V3 region of the 16S rRNA gene was sequenced. Results: 37.5% of CVS and 14% of AC samples showed the presence of bacterial DNA. Conclusion: Our study suggests that bacterial DNA can be identified in the placenta and amniotic fluid during early prenatal life.

Intrauterine versus post-mortem magnetic resonance in second trimester termination of pregnancy for central nervous system abnormalities.

OBJECTIVE: To evaluate if limiting factors of intrauterine magnetic resonance imaging (iuMRI) performed in the early second trimester of pregnancy (19-23 weeks) affect its accuracy in comparison to post-mortem MRI (pmMRI) in fetuses that underwent termination of pregnancy (TOP) for central nervous system (CNS) defects. STUDY DESIGN: This is a secondary analysis of a 10 years prospective observational study. Cases of TOP < 23 weeks for CNS malformation that had undergone neurosonography (NSG), iuMRI, pmMRI and autopsy were included. The agreement between iuMRI and pmMRI was calculated. The autopsy represented the gold-standard. RESULTS: Overall, 143 TOPs for fetal congenital anomaly underwent the post-mortem diagnostic protocol. Of these, 31 cases underwent iuMRI and pmMRI for CNS abnormality. Three cases were excluded due to brain autolysis at autopsy. Corpus callosum defects were the most represented (16/28; 57 %). In only one case of posterior fossa defect, pmMRI identified the presence of vermian hypoplasia not diagnosed at iuMRI. In 2 cases (7%), iuMRI added clinically relevant additional findings to NSG, that were posteriorly confirmed by pmMRI. CONCLUSIONS: The study shows that, at 19-23 weeks and for CNS defects, limiting factors that might influence the performance of iuMRI have little influence on iuMRI accuracy. This finding is particularly important for professionals who work in countries with legal bound for TOP in the early second trimester.

Identification of large-for-gestational age fetuses using antenatal customized fetal growth charts: Can we improve the prediction of abnormal labor course?

INTRODUCTION: Fetal overgrowth is an acknowledged risk factor for abnormal labor course and maternal and perinatal complications. The objective of this study was to evaluate whether the use of antenatal ultrasound-based customized fetal growth charts in fetuses at risk for large-for-gestational age (LGA) allows a better identification of cases undergoing caesarean section due to intrapartum dystocia. MATERIAL AND METHODS: An observational study involving four Italian tertiary centers was carried out. Women referred to a dedicated antenatal clinic between 35 and 38 weeks due to an increased risk of having an LGA fetus at birth were prospectively selected for the study purpose. The fetal measurements obtained and used for the estimation of the fetal size were biparietal diameter, head circumference, abdominal circumference and femur length, were prospectively collected. LGA fetuses were defined by estimated fetal weight (EFW) >95th centile either using the standard charts implemented by the World Health Organization (WHO) or the customized fetal growth charts previously published by our group. Patients scheduled for elective caesarean section (CS) or for elective induction for suspected fetal macrosomia or submitted to CS or vacuum extraction (VE) purely due to suspected intrapartum distress were excluded. The incidence of CS due to labor dystocia was compared between fetuses with EFW >95th centile according WHO or customized antenatal growth charts. RESULTS: Overall, 814 women were eligible, however 562 were considered for the data analysis following the evaluation of the exclusion criteria. Vaginal delivery occurred in 466 (82.9 %) women (435 (77.4 %) spontaneous vaginal delivery and 31 (5.5 %) VE) while 96 had CS. The EFW was >95th centile in 194 (34.5 %) fetuses according to WHO growth charts and in 190 (33.8 %) by customized growth charts, respectively. CS due to dystocia occurred in 43 (22.2 %) women with LGA fetuses defined by WHO curves and in 39 (20.5 %) women with LGA defined by customized growth charts (p 0.70). WHO curves showed 57 % sensitivity, 72 % specificity, 24 % PPV and 91 % NPV, while customized curves showed 52 % sensitivity, 73 % specificity, 23 % PPV and 91 % NPV for CS due to labor dystocia. CONCLUSIONS: The use of antenatal ultrasound-based customized growth charts does not allow a better identification of fetuses at risk of CS due to intrapartum dystocia.

Identification of Cannabis sativa L. (hemp) Retailers by Means of Multivariate Analysis of Cannabinoids.

In this work, the concentration of nine cannabinoids, six neutral cannabinoids (THC, CBD, CBC, CBG, CBN and CBDV) and three acidic cannabinoids (THCA CBGA and CBDA), was used to identify the Italian retailers of Cannabis sativa L. (hemp), reinforcing the idea that the practice of categorizing hemp samples only using THC and CBD is inadequate. A high-performance liquid chromatography/high-resolution mass spectrometry (HPLC-MS/MS) method was developed for screening and simultaneously analyzing the nine cannabinoids in 161 hemp samples sold by four retailers located in different Italian cities. The hemp samples dataset was analyzed by univariate and multivariate analysis with the aim to identify the hemp retailers without any other information on the hemp samples like Cannabis strains, seeds, soil and cultivation characteristics, geographical origin, product storage, etc. The univariate analysis highlighted that the hemp samples could not be differentiated by using any of the nine cannabinoids analyzed. To evaluate the real efficiency of the discrimination among the four hemp retailers a partial least squares discriminant analysis (PLS-DA) was applied. The PLS-DA results showed a very good discrimination between the four hemp retailers with an explained variance of 100% and low classification errors in both calibration (5%) and cross validation (6%). A total of 92% of the hemp samples were correctly classified by the cannabinoid variables in both fitting and cross validation. This work contributed to show that an analytical method coupled with multivariate analysis can be used as a powerful tool for forensic purposes.

CD56brightCD16(-) NK cells accumulate in psoriatic skin in response to CXCL10 and CCL5 and exacerbate skin inflammation.

Psoriasis is an immune-mediated skin disease characterized by lymphocytic infiltration and altered keratinocyte differentiation. Using immunohistochemical techniques we found that the cellular infiltrate in acute psoriatic plaques includes 5-8% CD3(-)CD56(+) natural killer (NK) cells, mostly localized in the mid and papillary dermis. NK lymphocytes isolated from punch biopsy specimens of psoriatic plaques showed a CD56(bright)CD16(-)CD158b(-) phenotype, failed to express the skin homing cutaneous lymphocyte-associated antigen and released abundant IFN-gamma upon stimulation. Supernatants from psoriatic NK cells induced MHC class II and ICAM-1 expression and release of CXCL10 and CCL5 by cultured psoriatic keratinocytes. Skin NK cells expressed high levels of the chemokines receptors CXCR3 and CCR5, intermediate amounts of CXCR1, CCR6 and CCR8, and low levels of CCR1, CCR2, CCR4, CCR7 and CX3CR1. In addition, they promptly migrated in vitro toward CXCL10, CCL5, supernatants of IFN-gamma-activated psoriatic keratinocytes and, to a lower extent, CCL20 and CCL4. In contrast, they failed to migrate toward CXCL8, CCL1, CCL2, CCL3, CCL17, CCL19 and CX3CL1. Taken together, our results implicate NK lymphocytes as newly identified protagonists in the pathogenesis of psoriasis. Their distinctive homing properties should be taken into account in the design of specific therapy aimed at blocking pathogenic cell accumulation in the skin.

Immunoregulation of allergic contact dermatitis.

Allergic contact dermatitis (ACD) to haptens can serve as a valuable paradigm for understanding the physiopathology of T cell mediated immune responses. In sensitized individuals, exposure to the relevant hapten initiates clinical expression of ACD, which depends on the rapid activation of specific T cells. Mechanisms of tissue damage include direct cytotoxicity against keratinocytes, mostly mediated by CD8+ T cells, and T cell release of cytokines, which amplify the inflammatory response by targeting resident skin cells. The expression of ACD is actively regulated by specialized subsets of T lymphocytes with suppressive functions. In particular, T regulatory cells producing high levels of IL-10 suppress ACD by blocking the functions of dendritic cells. In contrast CD4+CD25+ regulatory T cells prevent immunopathological reactions and maintain peripheral tolerance to haptens by acting via a cell-to-cell contact mechanism. Understanding the role of suppressor T cells and the requirements for their in vivo and in vitro expansion are critical steps for the development of specific desensitization protocols in hapten-allergic individuals. This information may also provide the basis for novel interventions in other immune-mediated diseases.

Human CD25+ regulatory T cells maintain immune tolerance to nickel in healthy, nonallergic individuals.

We investigated the capacity of CD25(+) T regulatory cells (Treg) to modulate T cell responses to nickel, a common cause of allergic contact dermatitis. CD4(+) T cells isolated from the peripheral blood of six healthy, nonallergic individuals showed a limited capacity to proliferate in response to nickel in vitro, but responsiveness was strongly augmented (mean increment +/- SD, 240 +/- 60%) when cells were depleted of CD25(+) Treg. Although CD25(+) Treg were anergic to nickel, a small percentage up-regulated membrane CTLA-4 upon nickel exposure. CD25(+) Treg strongly and dose-dependently inhibited nickel-specific activation of CD25(-) T lymphocytes in coculture experiments in a cytokine-independent, but cell-to-cell contact-dependent, manner. Approximately 30% of circulating CD25(+) Treg expressed the cutaneous lymphocyte-associated Ag (CLA), and CLA(+)CD25(+) Treg were more efficient than CLA(-)CD25(+) cells in suppressing nickel responsiveness of CD25(-) T cells. The site of a negative patch test in response to nickel showed an infiltrate of CD4(+)CLA(+) cells and CD25(+) cells, which accounted for approximately 20% of the total T cells isolated from the tissue. Skin-derived T cells suppressed nickel-specific responses of peripheral blood CD25(-) T cells. In addition, 60 +/- 14% of peripheral blood CD25(+) Treg expressed the chemokine receptor CCR7 and strongly inhibited naive T cell activation in response to nickel. Finally, CD25(+) T cells isolated from peripheral blood of nickel-allergic patients showed a limited or absent capacity to suppress metal-specific CD4(+) and CD8(+) T cell responses. The results indicates that in healthy individuals CD25(+) Treg can control the activation of both naive and effector nickel-specific T cells.

Immunoregulation of hapten and drug induced immune reactions.

PURPOSE OF REVIEW: Unbalanced immune responses to haptens lead to a variety of diseases, including allergic contact dermatitis to skin sensitizers and drug induced immune reactions. The occurrence, magnitude and persistence of immune responses are modulated by specialized T cell subsets with regulatory function. The purpose of this brief review is to summarize the recent data on the role of regulatory T cells in the control of hapten mediated diseases. RECENT FINDINGS: Several subsets of regulatory T cells, including T regulatory cell 1-like lymphocytes, and cutaneous lymphocyte associated antigen+ CD4+CD25+ regulatory T cells have been isolated from the skin at sites of hapten challenge. Both cell types suppress specific T cell responses to cutaneous sensitizers, such as nickel. SUMMARY: Although data concerning the regulation of drug hypersensitivity are lacking, several reports indicate the role of regulatory T cell subsets in allergic contact dermatitis to haptens. The understanding of their role in hapten diseases and the requirement for their in-vivo and in-vitro expansion appears as a critical step for the development of specific desensitization protocols.