Persistence of humoral response upon SARS-CoV-2 infection.

SARS-CoV-2 continues to leave its toll on global health and the economy. Management of the pandemic will rely heavily on the degree of adaptive immunity persistence following natural SARS-CoV-2 infection. Along with the progression of the pandemic, more literature on the persistence of the SARS-CoV-2-specific antibody response is becoming available. Here, we summarize findings on the persistence of the humoral, including neutralizing antibody, response at three to eight months post SARS-CoV-2 infection in non-pregnant adults. While the comparability of the literature is limited, findings on the detectability of immunoglobulin G class of antibodies (IgG) were most consistent and were reported in most studies to last for six to eight months. Studies investigating the response of immunoglobins M and A (IgM, IgA) were limited and reported mixed results, in particular, for IgM. The majority of studies observed neutralizing antibodies at all time points tested, which in some studies lasted up to eight months. The presence of neutralizing antibodies has been linked to protection from re-infection, suggesting long-term immunity to SARS-CoV-2. These neutralizing capacities may be challenged by emerging virus variants, but mucosal antibodies as well as memory B and T cells may optimize future immune responses. Thus, further longitudinal investigation of PCR-confirmed seropositive individuals using sensitive assays is warranted to elucidate the nature and duration of a more long-term humoral response.

Exhaled breath condensate as bioanalyte: from collection considerations to biomarker sensing.

Since the SARS-CoV-2 pandemic, the potential of exhaled breath (EB) to provide valuable information and insight into the health status of a person has been revisited. Mass spectrometry (MS) has gained increasing attention as a powerful analytical tool for clinical diagnostics of exhaled breath aerosols (EBA) and exhaled breath condensates (EBC) due to its high sensitivity and specificity. Although MS will continue to play an important role in biomarker discovery in EB, its use in clinical setting is rather limited. EB analysis is moving toward online sampling with portable, room temperature operable, and inexpensive point-of-care devices capable of real-time measurements. This transition is happening due to the availability of highly performing biosensors and the use of wearable EB collection tools, mostly in the form of face masks. This feature article will outline the last developments in the field, notably the novel ways of EBA and EBC collection and the analytical aspects of the collected samples. The inherit non-invasive character of the sample collection approach might open new doors for efficient ways for a fast, non-invasive, and better diagnosis.

[Pulmonary hypertension associated with left heart disease (group 2)]. / Pulmonale Hypertonie assoziiert mit Linksherzerkrankungen (Gruppe 2).

Pulmonary hypertension associated with left heart disease (PH-LHD) corresponds to group two of pulmonary hypertension according to clinical classification. Haemodynamically, this group includes isolated post-capillary pulmonary hypertension (IpcPH) and combined post- and pre-capillary pulmonary hypertension (CpcPH). PH-LHD is defined by an mPAP > 20 mmHg and a PAWP > 15 mmHg, pulmonary vascular resistance (PVR) with a cut-off value of 2 Wood Units (WU) is used to differentiate between IpcPH and CpcPH. A PVR greater than 5 WU indicates a dominant precapillary component. PH-LHD is the most common form of pulmonary hypertension, the leading cause being left heart failure with preserved (HFpEF) or reduced ejection fraction (HFmrEF, HFrEF), valvular heart disease and, less commonly, congenital heart disease. The presence of pulmonary hypertension is associated with increased symptom burden and poorer outcome across the spectrum of left heart disease. Differentiating between group 1 pulmonary hypertension with cardiac comorbidities and PH-LHD, especially due to HFpEF, is a particular challenge. Therapeutically, no general recommendation for the use of PDE5 inhibitors in HFpEF-associated CpcPH can be made at this time. There is currently no reliable rationale for the use of PAH drugs in IpcPH, nor is therapy with endothelin receptor antagonists or prostacyclin analogues recommended for all forms of PH-LHD.

Intracoronary autologous bone marrow cell transfer after myocardial infarction: the BOOST-2 randomised placebo-controlled clinical trial.

AIMS: Intracoronary infusion of autologous nucleated bone marrow cells (BMCs) enhanced the recovery of left ventricular ejection fraction (LVEF) after ST-segment elevation myocardial infarction (STEMI) in the randomised-controlled, open-label BOOST trial. We reassessed the therapeutic potential of nucleated BMCs in the randomised placebo-controlled, double-blind BOOST-2 trial conducted in 10 centres in Germany and Norway. METHODS AND RESULTS: Using a multiple arm design, we investigated the dose-response relationship and explored whether γ-irradiation which eliminates the clonogenic potential of stem and progenitor cells has an impact on BMC efficacy. Between 9 March 2006 and 16 July 2013, 153 patients with large STEMI were randomly assigned to receive a single intracoronary infusion of placebo (control group), high-dose (hi)BMCs, low-dose (lo)BMCs, irradiated hiBMCs, or irradiated loBMCs 8.1 ± 2.6 days after percutaneous coronary intervention (PCI) in addition to guideline-recommended medical treatment. Change in LVEF from baseline (before cell infusion) to 6 months as determined by MRI was the primary endpoint. The trial is registered at Current Controlled Trials (ISRCTN17457407). Baseline LVEF was 45.0 ± 8.5% in the overall population. At 6 months, LVEF had increased by 3.3 percentage points in the control group and 4.3 percentage points in the hiBMC group. The estimated treatment effect was 1.0 percentage points (95% confidence interval, -2.6 to 4.7; P = 0.57). The treatment effect of loBMCs was 0.5 percentage points (-3.0 to 4.1; P = 0.76). Likewise, irradiated BMCs did not have significant treatment effects. BMC transfer was safe and not associated with adverse clinical events. CONCLUSION: The BOOST-2 trial does not support the use of nucleated BMCs in patients with STEMI and moderately reduced LVEF treated according to current standards of early PCI and drug therapy.

Comparison of stroke volumes assessed by three-dimensional echocardiography and transpulmonary thermodilution in a pediatric animal model.

To compare stroke volumes (SV) in small hearts assessed by real-time three-dimensional echocardiography (3DE) with SV measured by transpulmonary thermodilution (TPTD) and continuous pulse contour analysis (PC) under various hemodynamic conditions. In thirteen anesthetized piglets (range 3.6-7.1 kg) SV were measured by 3DE, TPTD and PC at baseline and during phenylephrine and esmolol administration. 3DE and TPTD measurements were done successively while SV calculated by PC was documented at the time of 3DE. 3DE and TPTD showed a good correlation (r2 = 0.74) and a bias of -1.3 ml (limits of agreement -4.1 to 1.5 ml). While TPTD measured higher SV than 3DE, both methods tracked SV changes with a concordance rate of 91 %. PC and 3DE showed a lower correlation coefficient of r2 = 0.57 and a bias of -2.1 ml (limits of agreement -5.9 to 1.8 ml). Inter- and intra-observer variability of SV measured by 3DE was good with a mean bias <5 %. SV3DE showed a small variance and tracked acute small changes in SV in acceptable concordance with TPTD. PC measured SV with a higher variance and mean difference compared to 3DE. In an experimental setting 3DE has the possibility to offer non-invasive assessments of ventricular volumes volume changes. To determine whether 3DE could be used for SV assessment in a clinical routine our results need confirmation in a clinical setting.

Therapeutic efficacy of AAV1.SERCA2a in monocrotaline-induced pulmonary arterial hypertension.

BACKGROUND: Pulmonary arterial hypertension (PAH) is characterized by dysregulated proliferation of pulmonary artery smooth muscle cells leading to (mal)adaptive vascular remodeling. In the systemic circulation, vascular injury is associated with downregulation of sarcoplasmic reticulum Ca(2+)-ATPase 2a (SERCA2a) and alterations in Ca(2+) homeostasis in vascular smooth muscle cells that stimulate proliferation. We, therefore, hypothesized that downregulation of SERCA2a is permissive for pulmonary vascular remodeling and the development of PAH. METHODS AND RESULTS: SERCA2a expression was decreased significantly in remodeled pulmonary arteries from patients with PAH and the rat monocrotaline model of PAH in comparison with controls. In human pulmonary artery smooth muscle cells in vitro, SERCA2a overexpression by gene transfer decreased proliferation and migration significantly by inhibiting NFAT/STAT3. Overexpresion of SERCA2a in human pulmonary artery endothelial cells in vitro increased endothelial nitric oxide synthase expression and activation. In monocrotaline rats with established PAH, gene transfer of SERCA2a via intratracheal delivery of aerosolized adeno-associated virus serotype 1 (AAV1) carrying the human SERCA2a gene (AAV1.SERCA2a) decreased pulmonary artery pressure, vascular remodeling, right ventricular hypertrophy, and fibrosis in comparison with monocrotaline-PAH rats treated with a control AAV1 carrying ß-galactosidase or saline. In a prevention protocol, aerosolized AAV1.SERCA2a delivered at the time of monocrotaline administration limited adverse hemodynamic profiles and indices of pulmonary and cardiac remodeling in comparison with rats administered AAV1 carrying ß-galactosidase or saline. CONCLUSIONS: Downregulation of SERCA2a plays a critical role in modulating the vascular and right ventricular pathophenotype associated with PAH. Selective pulmonary SERCA2a gene transfer may offer benefit as a therapeutic intervention in PAH.

Characterizing preclinical models of ischemic heart failure: differences between LAD and LCx infarctions.

Large animal studies are an important step toward clinical translation of novel therapeutic approaches. We aimed to establish an ischemic heart failure (HF) model with a larger myocardial infarction (MI) relative to previous studies, and characterize the functional and structural features of this model. An MI was induced by occluding the proximal left anterior descending artery (LAD; n = 15) or the proximal left circumflex artery (LCx; n = 6) in Yorkshire pigs. Three pigs with sham procedures were also included. All pigs underwent hemodynamic and echocardiographic assessments before MI, at 1 mo, and 3 mo after MI. Analyses of left ventricular (LV) myocardial mechanics by means of strains and torsion were performed using speckle-tracking echocardiography and compared between the groups. The proximal LAD MI approach induced larger infarct sizes (14.2 ± 3.2% vs. 10.6 ± 1.9%, P = 0.03), depressed systolic function (LV ejection fraction; 39.8 ± 7.5% vs. 54.1 ± 4.6%, P < 0.001), and more LV remodeling (end-systolic volume index; 82 ± 25 ml/m(2) vs. 51 ± 18 ml/m(2), P = 0.02, LAD vs. LCx, respectively) compared with the LCx MI approach without compromising the survival rate. At the papillary muscle level, echocardiographic strain analysis revealed no differences in radial and circumferential strain between LAD and LCx MIs. However, in contrast with the LCx MI, the LAD MI resulted in significantly decreased longitudinal strain. The proximal LAD MI model induces more LV remodeling and depressed LV function relative to the LCx MI model. Location of MI significantly impacts the severity of HF, thus careful consideration is required when choosing an MI model for preclinical HF studies.

Inhibition of PKCα/ß with ruboxistaurin antagonizes heart failure in pigs after myocardial infarction injury.

RATIONALE: Protein kinase Cα (PKCα) activity and protein level are induced during cardiac disease where it controls myocardial contractility and propensity to heart failure in mice and rats. For example, mice lacking the gene for PKCα have enhanced cardiac contractility and reduced susceptibility to heart failure after long-term pressure overload or after myocardial infarction injury. Pharmacological inhibition of PKCα/ß with Ro-32-0432, Ro-31-8220 or ruboxistaurin (LY333531) similarly enhances cardiac function and antagonizes heart failure in multiple models of disease in both mice and rats. OBJECTIVE: Large and small mammals differ in several key indexes of heart function and biochemistry, lending uncertainty as to how PKCα/ß inhibition might affect or protect a large animal model of heart failure. METHODS AND RESULTS: We demonstrate that ruboxistaurin administration to a pig model of myocardial infarction-induced heart failure was protective. Twenty-kilogram pigs underwent left anterior descending artery occlusion resulting in myocardial infarctions and were then divided into vehicle or ruboxistaurin feed groups, after which they were monitored monthly for the next 3 months. Ruboxistaurin administered pigs showed significantly better recovery of myocardial contractility 3 months after infarction injury, greater ejection fraction, and greater cardiac output compared with vehicle-treated pigs. CONCLUSIONS: These results provide additional evidence in a large animal model of disease that PKCα/ß inhibition (with ruboxistaurin) represents a tenable and novel therapeutic approach for treating human heart failure.

Neutralizing antibodies against AAV serotypes 1, 2, 6, and 9 in sera of commonly used animal models.

Adeno-associated virus (AAV)-based vectors are promising gene delivery vehicles for human gene transfer. One significant obstacle to AAV-based gene therapy is the high prevalence of neutralizing antibodies in humans. Until now, it was thought that, except for nonhuman primates, pre-existing neutralizing antibodies are not a problem in small or large animal models for gene therapy. Here, we demonstrate that sera of several animal models of cardiovascular diseases harbor pre-existing antibodies against the cardiotropic AAV serotypes AAV1, AAV6, and AAV9 and against AAV2. The neutralizing antibody titers vary widely both between species and between serotypes. Of all species tested, rats displayed the lowest levels of neutralizing antibodies. Surprisingly, naive mice obtained directly from commercial vendors harbored neutralizing antibodies. Of the large animal models tested, the neutralization of AAV6 transduction by dog sera was especially pronounced. Sera of sheep and rabbits showed modest neutralization of AAV transduction whereas porcine sera strongly inhibited transduction by all AAV serotypes and displayed the largest variation between individual animals. Importantly, neutralizing antibody titers as low as 1/4 completely prevented in vivo transduction by AAV9 in rats. Our results suggest that prescreening of animals for neutralizing antibodies will be important for future gene transfer experiments in these animal models.

Concomitant intravenous nitroglycerin with intracoronary delivery of AAV1.SERCA2a enhances gene transfer in porcine hearts.

SERCA2a gene therapy improves contractile and energetic function of failing hearts and has been shown to be associated with benefits in clinical outcomes, symptoms, functional status, biomarkers, and cardiac structure in a phase 2 clinical trial. In an effort to enhance the efficiency and homogeneity of gene uptake in cardiac tissue, we examined the effects of nitroglycerin (NTG) in a porcine model following AAV1.SERCA2a gene delivery. Three groups of Göttingen minipigs were assessed: (i) group A: control intracoronary (IC) AAV1.SERCA2a (n = 6); (ii) group B: a single bolus IC injection of NTG (50 µg) immediately before administration of intravenous (IV) AAV1.SERCA2a (n = 6); and (iii) group C: continuous IV NTG (1 µg/kg/minute) during the 10 minutes of AAV1.SERCA2a infusion (n = 6). We found that simultaneous IV infusion of NTG and AAV1.SERCA2a resulted in increased viral transduction efficiency, both in terms of messenger RNA (mRNA) as well as SERCA2a protein levels in the whole left ventricle (LV) compared to control animals. On the other hand, IC NTG pretreatment did not result in enhanced gene transfer efficiency, mRNA or protein levels when compared to control animals. Importantly, the transgene expression was restricted to the heart tissue. In conclusion, we have demonstrated that IV infusion of NTG significantly improves cardiac gene transfer efficiency in porcine hearts.