Raman spectroscopy and imaging are highly structure-sensitive methods that allow the characterization of biological samples with minimal impact. In this paper, Raman spectra and imaging of ...noncancerous and cancerous human colon tissue samples were measured at different excitation wavelengths: 355, 532, and 785 nm. Intra-patient variability in the analyzed spectra showed colon sample heterogeneity for both noncancerous and cancerous human sample types. The lowest inter-patient variability of Raman spectra was observed for the fingerprint region of noncancerous samples for the 532 nm excitation laser line. The bands of principal biochemical constituents (proteins, lipids, nucleic acids) predominate in VIS and NIR-Raman spectra (excitation: 532, 785 nm), with the special role of the bands of intrinsic tissue chromophores-carotenoids for VIS excitation due to resonance enhancement. At 355 nm excitation, high autofluorescence of colon tissues were observed. Our studies proved high potential of Raman spectroscopy and Raman imaging in differentiation of noncancerous and cancerous human colon tissues and that the wavelengths 532 and 785 nm offer wide possibilities for the detection of human colon tissue pathology for
ex vivo
and
in vivo
measurements and prevail over 355 nm excitation.
Raman spectroscopy and imaging are highly structure-sensitive methods that allow the characterization of biological samples with minimal impact.
Background
Diagnosis of antiphospholipid syndrome (APS) requires persistent presence of lupus anticoagulant (LAC), anticardiolipin (aCL) IgG/IgM, or anti‐β2 glycoprotein I (aβ2GPI) IgG/IgM ...antibodies. Other antiphospholipid antibodies (aPL) such as antiphosphatidylserine/prothrombin antibodies (aPS/PT) are promising in assessment of thrombotic APS (TAPS).
Aim
To evaluate the added value of aPS/PT IgG and IgM in TAPS.
Material and Methods
aPS/PT IgG/IgM, aCL IgG/IgM, aβ2GPI IgG/IgM, and LAC were determined in 757 patients (TAPS and controls). aPS/PT cut‐off values were calculated, and aPS/PT titers and positivity were compared between TAPS and controls, type of thrombosis, and antibody profiles. Likelihood ratios (LR), odds ratios (OR), and aPL score were determined.
Results
aPS/PT IgG and IgM were associated with TAPS and triple positivity. In‐house calculated cut‐offs were higher for IgM (43 units), compared to manufacturer’s cut‐off (30 units). Thresholds of 90 (IgG) and 200 (IgM) units were determined as high‐titer cut‐off. Higher aPS/PT titers were observed in triple positive patients and showed higher LR and OR for TAPS. aPS/PT was independently associated with TAPS when adjusted for aCL/aβ2GPI, but not when adjusted for LAC. In isolated LAC positive patients, aPS/PT was positive in 27.1% TAPS patients and in 77.3% patients with autoimmune disease. Diagnostic value of aPL score did not differ with and without including aPS/PT.
Conclusion
aPS/PT positivity, especially with high antibody titer, is associated with TAPS diagnosis. Analysis on top of current laboratory criteria is not essential in TAPS diagnosis, but aPS/PT could be useful in patients with thrombosis and a double positive aPL profile (aCL+/aβ2GPI+).
Introduction
Granulomatosis with polyangiitis (GPA) is a small vessel vasculitis with a complex pathomechanism. Organ damage in GPA is also mediated by extracellular trap formation (NETosis). We ...analyzed the functional status of phosphoproteins modulating NETosis in neutrophils by the mammalian target of rapamycin (mTOR) pathway in GPA along with NETosis biomarkers.
Methods
Phosphoproteins levels measured in isolated neutrophils from 42 patients with GPA (exacerbation n=21; remission n=21) and 21 healthy controls were compared to serum biomarkers of the disease.
Results
Neutrophils in active disease manifested lowered levels of phosphorylated mTOR
(Ser2448),
PTEN
(Ser380)
and ULK1
(Ser555)
, whereas phosphorylated GSK-3α/β
(Ser21/Ser9)
was elevated. Exacerbation of GPA was characterized by elevated neutrophil dsDNA in serum, circulating mitochondrial DNA, and DNA-MPO complexes. A significant negative correlation between mTOR or PTEN phosphoproteins and biomarkers of GPA activity was also present, reflecting the clinical activity score of GPA. Positive correlations between phosphorylated GSK-3 α/β and circulating mtDNA, DNA-MPO complexes, neutrophil-released dsDNA, or circulating proteins were also significant. Increased serum levels of IGFBP-2, TFF-3, CD147, and CHI3L1 accompanied GPA exacerbation, whereas DPP-IV levels were the lowest in active GPA. Using a principal component analysis basigin, PTEN and mTOR had the highest loadings on the discrimination function, allowing classification between active, remission, and control subjects with 98% performance.
Conclusions
We present evidence that inhibited mTOR signaling accompanies NETosis in patients with GPA. The functional status of phosphoproteins suggests simultaneous activation of NETosis and autophagy. These results give rise to the study of autophagy as a mechanism underlying granuloma formation in GPA.
Abstract Antiphospholipid syndrome is an autoimmune disease which combines vascular thrombosis and/or pregnancy complications with the presence of antiphospholipid antibodies. It could be a ...devastating and sometimes life-threatening condition. As vascular thrombosis presents as typical venous or arterial thromboembolism diagnosis is based on laboratory data. Therefore proper performance and interpretation of laboratory tests is crucial. Broader knowledge about clinical and laboratory aspects of the syndrome and their associations are essential for proper evaluation and management of the patients.
Introduction: Immunoglobulin E is an important modulator of the inflammatory reaction in allergic asthma. It also contributes to airway remodeling in the course of the disease. The authors evaluated ...airway structural changes in severe allergic asthma during the omalizumab therapy. Patients and methods: The study included 13 patients with severe allergic asthma treated with omalizumab for at least one year. In each patient clinical, laboratory, and spirometry parameters were evaluated before and after the treatment. In addition, bronchoscopy with bronchial mucosa biopsy and bronchoalveolar lavage was performed. The basal lamina thickness, inflammatory cell infiltration, fibronectin, as well as type I and III collagen accumulation were assessed in bronchial mucosa specimens, together with the assessment of bronchoalveolar lavage cellularity. Results: The omalizumab therapy led to a decrease in the basal lamina thickness (p = 0.002), and to a reduction in fibronectin (p = 0.02), but not collagen deposits in the bronchial mucosa. The decrease in fibronectin accumulation was associated with an improvement in asthma control and quality of life (p = 0.01, both), and a diminished dose of systemic corticosteroids (p = 0.001). It was also associated with a tendency towards reduction of the eosinophil count in the peripheral blood, bronchoalveolar lavage fluid, and bronchial mucosa specimens. Conclusion: Our study has shown that omalizumab, effective in the treatment of severe allergic asthma, may also decrease unfavorable structural airway changes in allergic asthmatics, at least with respect to the fibronectin deposit and an increased thickness of the basal lamina. However, more extensive observational studies are needed to verify the above hypothesis.
The added value of antiphosphatidylserine/prothrombin antibodies (aPS/PT) in the diagnostic workup of antiphospholipid syndrome (APS) is unclear. Currently, diagnosis of thrombotic APS (TAPS) and ...obstetric APS (OAPS) requires persistent presence of lupus anticoagulant (LAC), anticardiolipin (aCL) immunoglobulin (Ig) G/IgM, or anti-β2-glycoprotein I (aβ2GPI) IgG/IgM antibodies.
To evaluate the role of aPS/PT IgG and IgM in OAPS.
aPS/PT IgG/IgM, aCL IgG/IgM, aβ2GPI IgG/IgM, and LAC were determined in 653 patients (OAPS, TAPS, and controls). In-house aPS/PT cut-off values were calculated, titers and prevalence were compared between OAPS, TAPS, and controls and type of pregnancy morbidity. Sensitivity, specificity, likelihood ratios, and odds ratios (OR) with 95% CI were calculated.
In OAPS, aPS/PT IgG and IgM showed an OR of 4.32 (95% CI, 2.54-7.36) and 3.37 (95% CI, 1.93-5.89), respectively, but the association was not independent of LAC. Prevalence and titers of aPS/PT IgG and IgM were lower in OAPS than in patients with TAPS. aPS/PT were more prevalent and showed higher titers in patients with late pregnancy loss than in patients with early pregnancy loss with a positivity of 86.4% and 39.3%, respectively. Higher aPS/PT titers did not increase the likelihood of having OAPS.
The added value of aPS/PT testing in the current diagnostic workup of OAPS seems limited compared with LAC, aCL, and aβ2GPI. aPS/PT might be useful in specific subsets of patients with OAPS. However, future multicentric studies are needed to elucidate the risk of less frequent and most severe obstetrical manifestations.
•Antiphosphatidylserine/prothrombin antibodies (aPS/PT) were investigated in obstetric antiphospholipid syndrome (OAPS).•Prevalence and titers of aPS/PT are lower in OAPS than in thrombotic APS.•aPS/PT prevalence and titers are higher in late pregnancy loss than in early pregnancy loss.•Added value of aPS/PT testing in OAPS seems limited compared with current diagnostic strategies.
The objective of our study was to evaluate the T‐helper (Th) and regulatory T (Treg) cell profile in ANCA‐positive granulomatosis with polyangiitis (GPA) and its relation to disease activity. In a ...prospective study, we studied two groups of GPA patients: (i) disease flare (active‐GPA, BVAS>6, n = 19), (ii) sustained remission (≥ 1‐year prior enrollment, inactive‐GPA, BVAS = 0, n = 18). 24 age‐sex matched healthy subjects served as controls. Active‐GPA patients were followed for 6 months and reevaluated during remission (early remission; n = 13). We analyzed subsets of Th‐cells (flow cytometry), production of signature cytokines by in vitro stimulated lymphocytes, and broad spectrum of serum cytokines (Luminex). In all GPA patients we observed expansion of effector Th17 cells, and increased production of IL‐17A by in vitro stimulated T cells, as compared to controls. Disease flare was characterized by marked reduction in Treg cells, whereas in sustained remission we showed expansion of both Treg and Th2 subset. Finally, analyzing the cytokine profile, we identified CCL23 and LIGHT, as potential biomarkers of active disease. We conclude that in GPA, expansion of Treg and Th2 lymphocytes in parallel to increased Th17 response is a characteristic feature of sustained remission. In contrast, Treg cells are markedly decreased in disease flare.
We evaluated functional subsets of T‐helper cells in granulomatosis with polyangiitis (GPA) patients. Disease flare was characterized by expansion of Th17 and marked reduction in Treg cells. We showed suppression of Th17 responses early after induction therapy, while sustained remission was linked with additional expansion of Treg and Th2 subsets.
Abstract Introduction Antiphospholipid syndrome (APS) is associated with the risk of both arterial and venous thrombosis. However, it is not known which factors might determine the location of ...thrombosis. Materials and Methods To retrospectively characterize factors associated with the risk of arterial thrombosis in a cohort of APS patients. Analysis included laboratory and clinical criteria of APS, together with classical cardiovascular risk factors and the possible role of platelet integrin α2 β1 (807 C/T) and αIIb β3 (PI A1/2) genetic polymorphisms. We enrolled 163 APS patients (123 women and 40 men aged 21-75; mean age 43 years); 78 suffered from arterial thrombosis. Results There were no significant differences in the frequency or titers of different antiphospholipid antibodies with the exception of slightly increased frequency of IgG anticardiolipin antibodies (ACL) in the arterial thrombosis group. Livedo reticularis was observed significantly more often in the arterial thrombosis group, particularly in stroke patients. In univariate analysis arterial thrombosis was associated with male gender (OR-2,201; p = 0,033), arterial hypertension (OR-2,81; p = 0,002) and hypercholesterolemia (OR-3,69; p = 0,001). On multivariate analysis arterial hypertension (OR = 1,78; p = 0,008) and hypercholesterolemia (OR = 2,001; p = 0,002) remained as independent risk factors for arterial thrombosis. Platelet glycoprotein polymorphisms studied did not show any significant associations with arterial thrombosis in APS patients. Conclusions Among APS patients those with ACL IgG antibodies, having livedo reticularis, and suffering from hypertension an hypercholesterolemia are at the increased risk of arterial thrombosis.
Background
Antiphosphatidylserine/prothrombin complex (aPS/PT) antibodies are recognized as a marker for antiphospholipid syndrome (APS). Dense and poorly lysable fibrin clots occur in thrombotic ...APS. Compact clots predict thromboembolism, but determinants of the unfavorable clot phenotype remain unknown in APS. We hypothesized that elevated aPS/PT antibodies determine unfavorable clot features.
Methods
In a cohort study involving 124 consecutive patients with thrombotic APS, we measured at baseline plasma fibrin clot permeability (Ks), efficiency of fibrinolysis (clot lysis time, CLT), and turbidity (off anticoagulation) along with immunoglobulin (Ig)G/IgM aPS/PT. During follow‐up, symptomatic thromboembolic events were recorded.
Results
Elevated IgG and IgM aPS/PT antibodies >30 international enzyme units (UI) were detected in 54.8% and 42.7% of APS patients, including 76.2% and 54% of lupus anticoagulant‐ (LA, n = 63) positive patients, respectively. Elevated IgG and IgM aPS/PT antibodies predicted low Ks (lower quartile, <6 × 10−9 cm2; odds ratio OR = 5.93, 95% confidence interval CI 2.09‐16.82 and OR = 11.79, 95% CI 4.10‐33.92) and prolonged CLT (top quartile, ≥116 min; OR = 4.85, 95% CI 2.42‐25.07 and OR = 6.04, 95% CI 2.42‐15.07). No such associations were observed for anticardiolipin or β2‐glycoprotein I antibodies or LA presence. During follow‐up (median 72.5, range 66‐83 months), thromboembolic events observed in 32 (26.7%, 4.6%/year) patients were independently predicted by IgG aPS/PT antibodies >30 UI (hazard ratio HR = 3.04, 95% CI 1.20‐8.88) and low Ks (HR = 3.00, 95% CI 1.41‐6.50).
Conclusions
We identified aPS/PT antibodies as a determinant of denser and poorly lysable plasma fibrin clot formation in APS patients. The association of elevated aPS/PT antibodies with thromboembolism in APS could be at least in part mediated by prothrombotic clot properties.
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