A Capillary-Force-Driven, Single-Cell Transfer Method for Studying Rare Cells.

The characterization of individual cells within heterogeneous populations (e.g., rare tumor cells in healthy blood cells) has a great impact on biomedical research. To investigate the properties of these specific cells, such as genetic biomarkers and/or phenotypic characteristics, methods are often developed for isolating rare cells among a large number of background cells before studying their genetic makeup and others. Prior to using real-world samples, these methods are often evaluated and validated by spiking cells of interest (e.g., tumor cells) into a sample matrix (e.g., healthy blood) as model samples. However, spiking tumor cells at extremely low concentrations is challenging in a standard laboratory setting. People often circumvent the problem by diluting a solution of high-concentration cells, but the concentration becomes inaccurate after series dilution due to the fact that a cell suspension solution can be inhomogeneous, especially when the cell concentration is very low. We report on an alternative method for low-cost, accurate, and reproducible low-concentration cell spiking without the use of external pumping systems. By inducing a capillary force from sudden pressure drops, a small portion of the cellular membrane was aspirated into the reservoir tip, allowing for non-destructive single-cell transfer. We investigated the surface membrane tensions induced by cellular aspiration and studied a range of tip/tumor cell diameter combinations, ensuring that our method does not affect cell viability. In addition, we performed single-cell capture and transfer control experiments using human acute lymphoblastic leukemia cells (CCRF-CEM) to develop calibrated data for the general production of low-concentration samples. Finally, we performed affinity-based tumor cell isolation using this method to generate accurate concentrations ranging from 1 to 15 cells/mL.

Comparison of sample preparation methods for rare cell isolation in microfluidic devices.

The analysis of circulating tumor cells (CTCs) is important for cancer diagnosis and prognosis. Microfluidics has been employed for CTC analysis due to its scaling advantages and high performance. However, pre-analytical methods for CTC sample preparation are often combined with microfluidic platforms because a large sample volume is required to detect extremely rare CTCs. Among pre-analytical methods, Ficoll-Paque™, OncoQuick™, and RosetteSep™ are commonly used to separate cells of interest. To compare their performance, we spiked L3.6pl pancreatic cancer cells into healthy blood samples and then employed each technique to prepare blood samples, followed by using a microfluidic platform to capture and detect L3.6pl cells. We found these three methods have similar performance, though the slight edge of RosetteSep™ over Ficoll-Paque™ is statistically significant. We also studied the effects of the tumor cell concentrations on the performance of the frequently used Ficoll-Paque™ method. Furthermore, we examined the repeatability and variability of each pre-analytical technique and the microfluidics-enabled detection. This study will provide researchers and clinicians with comparative data that can influence the choice of sample preparation method, help estimate CTC loss in each pre-analytical method, and correlate the results of clinical studies that employ different techniques.

L'analyse des cellules tumorales circulantes (CTC) est une pierre angulaire du diagnostic et du pronostic du cancer. On recourt à la microfluidique pour l'analyse des CTC en raison des avantages qu'elle offre pour la mise à l'échelle et de sa grande performance. Par ailleurs, les méthodes préanalytiques pour la préparation d'échantillons de CTC font souvent appel à des plateformes microfluidiques, car il faut un grand volume d'échantillon pour détecter des CTC extrêmement rares. Parmi les méthodes préanalytiques couramment utilisées pour séparer les cellules sanguines d'intérêt, notons Ficoll-PaqueMC, OncoQuickMC et RosetteSepMC. Afin de comparer les performances de ces méthodes, nous avons additionné de cellules de cancer du pancreas L3,6pl des échantillons de sang sains, puis nous avons utilisé les trois méthodes pour préparer les échantillons sanguins, que nous avons ensuite soumis à une plateforme microfluidique pour isoler et détecter les cellules L3,6pl. Nos résultats montrent que les performances de ces trois méthodes sont similaires, bien que le léger avantage de RosetteSepMC par rapport à Ficoll-PaqueMC soit statistiquement significatif. Nous avons également étudié les effets des concentrations de cellules tumorales sur la performance de la méthode Ficoll-PaqueMC, qui est la plus fréquemment utilisée. En outre, nous avons examiné la répétabilité et la variabilité de chaque méthode préanalytique et les caractéristiques de détection que permet d'obtenir la microfluidique. Cette étude fournit aux chercheurs et aux cliniciens des données comparatives qui peuvent influencer leur choix de la méthode de préparation des échantillons, et permet d'estimer la perte de CTC propre à chaque méthode préanalytique et de corréler les résultats des études cliniques qui utilisent différentes techniques. [Traduit par la Rédaction].

Clinical and immunological study of Tofacitinib and Baricitinib in refractory Blau syndrome: case report and literature review.

Blau syndrome (BS) is an autoinflammatory disorder characterized by non-caseating granulomatous dermatitis, arthritis, and uveitis. We present a case of refractory and severe BS that was treated with the Janus kinase inhibitors (JAKINIBS), Tofacitinib (TOFA) and then Baricitinib (BARI). Our aim was to describe the clinical and immunological outcomes after treatment with JAKINIBS. Blood tests and serum samples were obtained during follow-up with TOFA and BARI. We assessed their effects on clinical outcomes, acute phase reactants, absolute lymphocyte counts (ALCs), lymphocyte subset counts, immunoglobulins, and cytokine levels. A review of the literature on the use of JAKINIBS for the treatment of uveitis and sarcoidosis was also conducted. TOFA led to a rapid and maintained disease control and a steroid-sparing effect. A decrease from baseline was observed in ALC, CD3+, CD4+, CD8+, and natural killer (NK) cell counts. B-cells were stable. Serum levels of interleukin (IL)-4 and tumor necrosis factor alpha (TNF-α) increased, whereas IL-2, IL-6, IL-10, and IL-17 maintained stable. TOFA was discontinued after 19 months due to significant lymphopenia. The initiation of BARI allowed maintaining adequate control of disease activity with an adequate safety profile. The literature review showed seven patients with uveitis and five with sarcoidosis treated with JAKINIBS. No cases of BS treated with JAKINIBS were found. We report the successful use of JAKINIBS in a patient with refractory and severe BS.

Intravenous methylprednisolone induces rapid improvement in non-infectious uveitis: a multicentre study of 112 patients.

OBJECTIVES: Rapid control of intraocular inflammation in non-infectious uveitis (NIU) is mandatory to avoid irreversible structural and functional damage. In this study, we assessed the efficacy and safety of intravenous methylprednisolone (IVMP) pulses in the treatment of NIU. METHODS: A retrospective case series of 112 patients who received IVMP for the treatment of NIU, either isolated or associated with different underlying diseases, was studied. Intraocular inflammation (anterior chamber cells and vitritis) was the primary outcome measure. Secondary outcome measures were macular thickness and best corrected visual acuity (BCVA). Patients were assessed at baseline visit, and at days 2-5, 7, 15 and 30 after initiation of IVMP pulse therapy. RESULTS: A total of 112 patients (mean age 42±14.5 yrs) were assessed. An underlying immune-mediated disease was diagnosed in 73 patients. Inflammatory ocular patterns were panuveitis (n=68), posterior uveitis (n=30), anterior uveitis (AU) (n=12), and intermediate uveitis (n=2). Additionally, patients presented cystoid macular oedema (CME) (n=50), retinal vasculitis (n=37), and exudative retinal detachment (n=31). Therapies used before IVMP included intraocular glucocorticoids (n=4), high-dose oral systemic glucocorticoids (n=77), and conventional (n=107) or biologic (n=40) immunosuppressive drugs. IVMP dose ranged from 80 to 1,000 mg/day for 3-5 consecutive days. Improvement was observed in AU, vitritis, BCVA, CME, and retinal vasculitis. At first month evaluation, total remission was achieved in 19 patients. Side effects of IVMP were respiratory infections (n=3), uncontrolled hyperglycaemia (n=1), herpes zoster (n=1), and oral candidiasis (n=1). CONCLUSIONS: IVMP pulse therapy was effective and safe, and achieved rapid control of NIU.

Epidemiology of sarcoidosis in northern Spain, 1999-2019: A population-based study.

BACKGROUND: The incidence of sarcoidosis varies widely worldwide. The aim of this study was to estimate the incidence of sarcoidosis in a population-based cohort from northern Spain. METHODS: Patients diagnosed with sarcoidosis at Marqués de Valdecilla University Hospital, corresponding to the central Cantabria that encompasses Santander city and the surroundings, between January 1999 and December 2019were assessed. The diagnosis of sarcoidosis was established according to ATS/ERS/WASOG criteria as follows: compatible clinical and radiological presentation, histopathologic confirmation, and exclusion of other granulomatous diseases. Demographic and clinical data were collected. The incidence of sarcoidosis between 1999-2019 was estimated by sex, age, and year of diagnosis. RESULTS: A total of 234 patients were included, with a male/female ratio of 0.81. The mean age of the cohort at diagnosis was 48.43 ± 14.83 years and 129 (55.1%) were women. Incidence during the period of study was 3.58 per 100,000 populations (95% confidence interval: 3.13 - 4.07). No gender predominance was observed. An increase in age at diagnosis over time was found in the linear regression analysis. Thoracic affection was found in 180 patients (76.9%). Most common extra-thoracic areas affected were skin (34.2%), joints (30.8%) and eyes (15.4%). CONCLUSIONS: The incidence of sarcoidosis estimated in this study was similar to that of other Mediterranean countries. No gender predominance was observed. Consistent with previous studies, male presented an incidence peak 10 years earlier than female. A second peak between ages 60-69 years was identified in both sexes.

Treatment With Tofacitinib in Refractory Psoriatic Arthritis: A National Multicenter Study of the First 87 Patients in Clinical Practice.

OBJECTIVE: Tofacitinib (TOF) is the first Janus kinase (JAK) inhibitor approved for psoriatic arthritis (PsA). It has shown efficacy in patients refractory to anti-tumor necrosis factor-α in randomized controlled trials (RCTs). Our aim was to assess efficacy and safety of TOF in clinical practice. METHODS: This was an observational, open-label multicenter study of PsA patients treated with TOF due to inefficacy or adverse events of previous therapies. Outcome variables were efficacy, corticosteroid dose-sparing effect, retention rate, and safety. A comparative study of clinical features between our cohort of patients and those from the OPAL Beyond trial was performed. RESULTS: There were 87 patients (28 women/59 men), with a mean age of 52.8 ± 11.4 years. All patients were refractory to biologic disease-modifying antirheumatic drugs (DMARDs) and/or to conventional synthetic DMARDs plus apremilast. TOF was started at 5 mg twice daily after a mean follow-up of 12.3 ± 9.3 years from PsA diagnosis. At first month, Disease Activity Score in 28 joints based on erythrocyte sedimentation rate (DAS28-ESR) decreased from median 4.8 (IQR 4.1-5.4) to 3.7 (IQR 2.8-4.7, P < 0.01), Disease Activity Index for Psoriatic Arthritis from median 28 (IQR 18.4-34.1) to 15.5 (IQR 10.1-25.7, P < 0.01), and C-reactive protein from median 1.9 (IQR 0.3-5.0) to 0.5 (IQR 0.1-2.2) mg/dL (P < 0.01). Also, TOF led to a significant reduction in prednisone dose. Mild adverse effects were reported in 21 patients (24.13%), mainly gastrointestinal symptoms. TOF retention rate at Month 6 was 77% (95% CI 65.2-86.3). Patients in clinical practice were older with longer disease duration and received biologic agents more commonly than those in the OPAL Beyond trial. CONCLUSION: Data from clinical practice confirm that TOF seems to be effective, rapid, and relatively safe in refractory PsA despite clinical differences with patients in RCTs.

Immune-related adverse events in patients with solid-organ tumours treated with immunotherapy: a 3-year study of 102 cases from a single centre.

OBJECTIVES: Immune checkpoint blockade therapy (ICBT) increases the anti-tumoural function of the immune system, but it can also induce immune-related adverse events (irAEs). Our aim was to assess the irAEs due to ICBT in patients from a single centre of Northern Spain. METHODS: We set up an observational study of patients treated in monotherapy with ICBT targeted against cytotoxic T-lymphocyte antigen 4 (CTLA-4), programmed cell death 1 (PD-1) or its ligand (PD-L1) for solid organ tumours. All patients were followed up in a single University Hospital from March 2015 to September 2018. RESULTS: We studied 102 patients (63 men/39 women); mean age 60.6±9.7 years, with lung (n=63), melanoma (n=21), kidney (n=11), gastric (n=3), colon (n=3) or bladder (n=1) cancer. Only 7 patients had a previous diagnosis of an immune-mediated disease, specifically: psoriasis (n=2), psoriatic arthritis (n=1), systemic lupus erythematosus (n=1), spondyloarthitis (n=1), rheumatoid arthritis (n=1) and cutaneous lupus (n=1). One of the following ICBT was administered: nivolumab (n=52), pembrolizumab (n=35), atezolizumab (n=10) and ipilimumab (n=5). After a mean follow-up time of 14.4±7.7 months since ICBT onset, 87 (85.3%) patients had experienced irAEs, mostly gastrointestinal, thyroid and musculskeletal manifestations including inflammatory arthralgia (n= 8), arthritis (n= 6) and myositis (n=2). ICBT was discontinued in 41 patients but it was reintroduced in 30 of them after resolution of the adverse event, with a good tolerance in all cases. Thirty-six (41.4%) of the 87 patients required specific treatment (prednisone, levothyroxine, and thiamazol) for the irAEs. CONCLUSIONS: irAEs are frequent in patients undergoing ICBT. Almost half of the patients that have irAEs require treatment. Musculoskeletal manifestations are not uncommon.

Long-term Follow-up and Optimization of Infliximab in Refractory Uveitis Due to Behçet Disease: National Study of 103 White Patients.

OBJECTIVE: In a large series of White patients with refractory uveitis due to Behçet disease (BD) being treated with infliximab (IFX), we assessed (1) long-term efficacy and safety of IFX, and (2) IFX optimization when ocular remission was achieved. METHODS: Our multicenter study of IFX-treated patients with BD uveitis refractory to conventional immunosuppressant agents treated 103 patients/185 affected eyes with IFX as first biologic therapy in the following intervals: 3-5 mg/kg intravenous at 0, 2, 6, and then every 4-8 weeks. The main outcome variables were analyzed at baseline, first week, first month, sixth month, first year, and second year of IFX therapy. After remission, based on a shared decision between patient and clinician, IFX optimization was performed. Efficacy, safety, and cost of IFX therapy were evaluated. RESULTS: In the whole series (n = 103), main outcome variables showed a rapid and maintained improvement, reaching remission in 78 patients after a mean IFX duration of 31.5 months. Serious adverse events were observed in 9 patients: infusion reactions (n = 4), tuberculosis (n = 1), Mycobacterium avium pneumonia (n = 1), severe oral ulcers (n = 1), palmoplantar psoriasis (n = 1), and colon carcinoma (n = 1). In the optimization subanalysis, the comparative study between optimized and nonoptimized groups showed (1) no differences in clinical characteristics at baseline, (2) similar maintained improvement in most ocular outcomes, (3) lower severe adverse events, and (4) lower mean IFX costs in the optimized group (€4826.52 vs €9854.13 per patient/yr). CONCLUSION: IFX seems to be effective and relatively safe in White patients with refractory BD uveitis. IFX optimization is effective, safe, and cost-effective.

Incorporation of lateral microfiltration with immunoaffinity for enhancing the capture efficiency of rare cells.

The methods for isolating rare cells such as circulating tumor cells (CTCs) can be generally classified into two categories: those based on physical properties (e.g., size) and methods based on biological properties (e.g., immunoaffinity). CellSearch, the only FDA-approved method for the CTC-based cancer prognosis, relies on immunoaffinity interactions between CTCs and antibodies immobilized on magnetic particles. Immunoaffinity-based CTC isolation has also been employed in microfluidic devices, which show higher capture efficiency than CellSearch. We report here our investigation of combining size-based microfiltration into a microfluidic device with immunoaffinity for enhanced capture efficiency of CTCs. The device consists of four serpentine main channels, and each channel contains an array of lateral filters that create a two-dimensional flow. The main flow is through the serpentine channel, allowing the majority of the sample to pass by while the secondary flow goes through the lateral filters. The device design is optimized to make all fluid particles interact with filters. The filter sizes range from 24 to 12 µm, being slightly larger than or having similar dimension of CTCs. These filters are immobilized with antibodies specific to CTCs and thus they function as gates, allowing normal blood cells to pass by while forcing the interactions between CTCs and antibodies on the filter surfaces. The hydrodynamic force experienced by a CTC was also studied for optimal experimental conditions to ensure immunoaffinity-enabled cell capture. The device was evaluated by capturing two types of tumor cells spiked in healthy blood or a buffer, and we found that their capture efficiency was between 87.2 and 93.5%. The platform was further validated by isolating CTCs from blood samples of patients with metastatic pancreatic cancer.

Integration of Lateral Filter Arrays with Immunoaffinity for Circulating-Tumor-Cell Isolation.

Circulating tumor cells (CTCs) are an important biomarker for cancer prognosis and treatment monitoring. However, the heterogeneity of the physical and biological properties of CTCs limits the efficiency of various approaches used to isolate small numbers of CTCs from billions of normal blood cells. To address this challenge, we developed a lateral filter array microfluidic (LFAM) device to integrate size-based separation with immunoaffinity-based CTC isolation. The LFAM device consists of a serpentine main channel, through which most of a sample passes, and an array of lateral filters for CTC isolation. The unique device design produces a two-dimensional flow, which reduces nonspecific, geometric capture of normal cells as typically observed in vertical filters. The LFAM device was further functionalized by immobilizing antibodies that are specific to the target cells. The resulting devices captured pancreatic cancer cells spiked in blood samples with (98.7±1.2) % efficiency and were used to isolate CTCs from patients with metastatic colorectal cancer.

Microfluidic Isolation of Circulating Tumor Cells and Cancer Stem-Like Cells from Patients with Pancreatic Ductal Adenocarcinoma.

Background: Pancreatic ductal adenocarcinoma (PDAC) requires multimodal therapeutic approaches and disease monitoring for effective treatment. Liquid biopsy biomarkers, including circulating tumor cells (CTCs) and cancer stem-like cells (CSCs), hold promise for evaluating treatment response promptly and guiding therapeutic modifications. Methods: From 24 patients with metastatic PDAC (stage IV, M1) undergoing active systemic treatment, we collected 78 blood samples at different time points for CTC and CSC isolation using a microfluidic platform functionalized with antibodies against a CTC biomarker, epithelial cell adhesion molecule (EpCAM), or a CSC biomarker, CD133. These isolated cells were further verified, via fluorescent staining and imaging, using cytokeratin (CK), CD45, and nucleic acid stain 4',6-diamidino-2-phenylindole (DAPI). Results: The majority (84.4%) of patient blood samples were positive for CTCs (EpCAM+CK+CD45-DAPI+) and 70.8% of patient blood samples were positive for CSCs (CD133+CK+CD45-DAPI+), using the highest baseline value of healthy samples as threshold. The CTC subtypes (EpCAM+CK+CD45-DAPI+CD133+ and EpCAM+CK+CD45-DAPI+CD133-) and CSC subtypes (CD133+CK+CD45-DAPI+EpCAM+ and CD133+CK+CD45-DAPI+EpCAM-) were also analyzed using immunochemical methods. In several cases, CSCs exhibited cytokeratin expression that did not express EpCAM, indicating that they will not be detected using EpCAM-based isolation. Conclusion: The microfluidic platform enabled the reliable isolation of CTCs and CSCs from PDAC patient samples, as well as their subtypes. Complementary assessment of both CTCs and CSCs appears advantageous to assess the profile of tumor progressing in some cases. This research has important implications for the application and interpretation of approved methods to detect CTCs.

Anti-IL6-Receptor Tocilizumab in Refractory and Noninfectious Uveitic Cystoid Macular Edema: Multicenter Study of 25 Patients.

PURPOSE: Cystoid macular edema (CME) is a leading cause of blindness. This study assessed the efficacy and safety of tocilizumab (TCZ) in refractory CME. DESIGN: Retrospective case series. METHODS: Patients with CME secondary to noninfectious uveitis who had inadequate response to corticosteroids and at least 1 conventional immunosuppressive drug, and in most cases to other biological agents, were studied. CME was defined as central retinal thickness greater than 300 µm. The primary outcome measure was macular thickness. Intraocular inflammation, best-corrected visual acuity (BCVA), and corticosteroid-sparing effect were also analyzed. RESULTS: A total of 25 patients (mean ± standard deviation age 33.6 ± 18.9 years; 17 women) with CME were assessed. Underlying diseases associated with uveitis-related CME are juvenile idiopathic arthritis (n = 9), Behçet disease (n = 7), birdshot retinochoroidopathy (n = 4), idiopathic (n = 4), and sarcoidosis (n = 1). The ocular patterns were panuveitis (n = 9), anterior uveitis (n = 7), posterior uveitis (n = 5), and intermediate uveitis (n = 4). Most patients had CME in both eyes (n = 24). TCZ was used in monotherapy (n = 11) or combined with conventional immunosuppressive drugs. Regardless of the underlying disease, compared to baseline, a statistically significant improvement in macular thickness (415.7 ± 177.2 vs 259.1 ± 499.5 µm; P = .00009) and BCVA (0.39 ± 0.31 vs 0.54 ± 0.33; P = .0002) was obtained, allowing us to reduce the daily dose of prednisone (15.9 ± 13.6 mg/day vs 3.1 ± 2.3 mg/day; P = .002) after 12 months of therapy. Remission was achieved in 14 patients. Only minor side effects were observed after a mean follow-up of 12.7 ± 8.34 months. CONCLUSION: Macular thickness is reduced following administration of TCZ in refractory uveitis-related CME.

Successful Optimization of Adalimumab Therapy in Refractory Uveitis Due to Behçet's Disease.

PURPOSE: To assess efficacy, safety, and cost-effectiveness of adalimumab (ADA) therapy optimization in a large series of patients with uveitis due to Behçet disease (BD) who achieved remission after the use of this biologic agent. DESIGN: Open-label multicenter study of ADA-treated patients with BD uveitis refractory to conventional immunosuppressants. SUBJECTS: Sixty-five of 74 patients with uveitis due to BD, who achieved remission after a median ADA duration of 6 (range, 3-12) months. ADA was optimized in 23 (35.4%) of them. This biologic agent was maintained at a dose of 40 mg/subcutaneously/2 weeks in the remaining 42 patients. METHODS: After remission, based on a shared decision between the patient and the treating physician, ADA was optimized. When agreement between patient and physician was reached, optimization was performed by prolonging the ADA dosing interval progressively. Comparison between optimized and nonoptimized patients was performed. MAIN OUTCOME MEASURES: Efficacy, safety, and cost-effectiveness in optimized and nonoptimized groups. To determine efficacy, intraocular inflammation (anterior chamber cells, vitritis, and retinal vasculitis), macular thickness, visual acuity, and the sparing effect of glucocorticoids were assessed. RESULTS: No demographic or ocular differences were found at the time of ADA onset between the optimized and the nonoptimized groups. Most ocular outcomes were similar after a mean ± standard deviation follow-up of 34.7±13.3 and 26±21.3 months in the optimized and nonoptimized groups, respectively. However, relevant adverse effects were only seen in the nonoptimized group (lymphoma, pneumonia, severe local reaction at the injection site, and bacteremia by Escherichia coli, 1 each). Moreover, the mean ADA treatment costs were lower in the optimized group than in the nonoptimized group (6101.25 euros/patient/year vs. 12 339.48; P < 0.01). CONCLUSION: ADA optimization in BD uveitis refractory to conventional therapy is effective, safe, and cost-effective.

Anti-interleukin 6 receptor tocilizumab in refractory uveitis associated with Behçet's disease: multicentre retrospective study.

Objective: To assess the efficacy of tocilizumab (TCZ) in refractory uveitis of Behçet's disease (BD). Methods: Multicentre study of patients with BD-associated uveitis. Patients were refractory to conventional and biologic immunosuppressive drugs. The main outcome measures were intraocular inflammation, macular thickness, visual acuity and corticosteroid-sparing effects. Results: We studied 11 patients (7 men) (20 affected eyes); median age 35 years. Uveitis was bilateral in nine patients. The patterns of ocular involvement were panuveitis (n = 8, with retinal vasculitis in 4), anterior uveitis (n = 2) and posterior uveitis (n = 1). Cystoid macular oedema was present in seven patients. The clinical course was recurrent (n = 7) or chronic (n = 4). Before TCZ, patients had received systemic corticosteroids, conventional immunosuppressants and the following biologic agents: adalimumab (n = 8), infliximab (n = 4), canakimumab (n = 1), golimumab (n = 3), etanercept (n = 1). TCZ was used as monotherapy or combined with conventional immunosuppressants at 8 mg/kg/i.v./4 weeks (n = 10) or 162 mg/s.c./week (n = 1). At TCZ onset the following extraocular manifestations were present: oral and/or genital ulcers (n = 7), arthritis (n = 4), folliculitis/pseudofolliculitis (n = 4), erythema nodosum (n = 2), livedo reticularis (n = 1) and neurological involvement (n = 2). TCZ yielded rapid and maintained improvement in all ocular parameters of the patients, with complete remission in eight of them. However, this was not the case for the extraocular manifestations, since TCZ was only effective in three of them. After a mean (s.d.) follow-up of 9.5 (8.05) months, TCZ was withdrawn in two cases, due to a severe infusion reaction and arthritis impairment, respectively. Conclusion: TCZ could be a therapeutic option in patients with BD and refractory uveitis.

Golimumab in refractory uveitis associated to juvenile idiopathic arthritis: multicentre study of 7 cases and literature review.

OBJECTIVES: To assess the efficacy of golimumab (GLM), a fully humanised anti-TNF-α monoclonal antibody, in refractory juvenile idiopathic arthritis (JIA)-associated uveitis. METHODS: This was a multicentre study of JIA-associated uveitis refractory to standard synthetic immunosuppressive drugs and in most cases to other anti-TNF-α agents. Results were expressed as mean±standard deviation or as median (range or interquartile range). The Wilcoxon signed-rank test was used to compare continuous variables. A literature review of the efficacy of GLM in uveitis related to JIA was also conducted. RESULTS: We studied 7 patients (5 females; mean age 21.7±7.5 years; 13 affected eyes). Uveitis was bilateral in 6. Cystoid macular oedema (CME) occurred in 3 patients (5 eyes). Besides corticosteroids and synthetic immunosuppressive drugs, patients had received before GLM a median of 2 biologic agents (range 0-3) including adalimumab (n=6), etanercept (n=2), infliximab (n=3) and abatacept (n=2). GLM dose was 50 mg/sc every 4 weeks. After 6 months of therapy the number of anterior chamber cells decreased from 1 [0.25-1.5] to 0 [0-0.5] (p=0.02) and optical coherence tomography (in patients with CME) from 313.6±77.05 to 261.4±75.1 µm (p=0.03). The best-corrected visual acuity increased from 0.5 to 0.62 (p=0.018). Complete remission of uveitis was achieved in 4 of 7 patients after 16.8±11.4 months of follow-up. However, 2 of the seven patients had to be switched to tocilizumab due to inefficacy. Local erythema at the injection site was observed in 2. CONCLUSIONS: GLM may be considered in the management of refractory JIA-related uveitis.

Tumor cell capture patterns around aptamer-immobilized microposts in microfluidic devices.

Circulating tumor cells (CTCs) have shown potential for cancer diagnosis and prognosis. Affinity-based CTC isolation methods have been proved to be efficient for CTC detection in clinical blood samples. One of the popular choices for affinity-based CTC isolation is to immobilize capture agents onto an array of microposts in microchannels, providing high CTC capture efficiency due to enhanced interactions between tumor cells and capture agents on the microposts. However, how the cells interact with microposts under different flow conditions and what kind of capture pattern results from the interactions have not been fully investigated; a full understanding of these interactions will help to design devices and choose experimental conditions for higher CTC capture effeciency. We report our study on their interaction and cell distribution patterns around microposts under different flow conditions. Human acute lymphoblastic leukemia cells (CCRF-CEM) were used as target cancer cells in this study, while the Sgc8 aptamer that has specific binding with CCRF-CEM cells was employed as a capture agent. We investigated the effects of flow rates and micropost shapes on the cell capture efficiency and capture patterns on microposts. While a higher flow rate decreased cell capture efficiency, we found that the capture pattern around microposts also changed, with much more cells captured in the front half of a micropost than at the back half. We also found the ratio of cells captured on microposts to the cells captured by both microposts and channel walls increased as a function of the flow rate. We compared circular microposts with an elliptical shape and found that the geometry affected the capture distribution around microposts. In addition, we have developed a theoretical model to simulate the interactions between tumor cells and micropost surfaces, and the simulation results are in agreement with our experimental observation.

A Novel Microfluidic Device for Isolation of Circulating Tumor Cells from Pancreatic Cancer Blood Samples.

Enumeration of circulating tumor cells (CTCs) can provide valuable prognostic information to guide cancer treatment as well as help monitor disease progression. Analysis of these rare malignant cells has the potential to further our understanding of cancer metastasis by gaining insights into CTC characteristics and properties. Microfluidics presents a unique platform to isolate and study CTCs. In this chapter, we describe the detailed procedures for the fabrication and use of a microfluidic device to detect CTCs from the blood of pancreatic cancer patients.

Significant sE-Selectin levels reduction after 6 months of anti-TNF-α therapy in non-diabetic patients with moderate-to-severe psoriasis.

PURPOSE: Psoriasis patients have high risk of atherosclerosis, characterized by endothelial dysfunction. We aimed to study the association of the endothelial activation biomarkers monocyte chemoattractant protein 1 (MCP-1), soluble (s) E-selectin and P-selectin with disease activity and severity in psoriasis patients treated with anti-TNF-α therapy. Also, to evaluate the relationship of metabolic syndrome features with these biomarkers and the effect of anti-TNF-α therapy on these molecules. METHODS: Twenty-nine consecutive non-diabetic patients with moderate-to-severe psoriasis who underwent 6 months of anti-TNF-α-adalimumab therapy were studied. Metabolic and clinical evaluation was performed prior to anti-TNF-α treatment (time 0) and 6 months later. MCP-1, sE-selectin and sP-selectin serum levels were determined by ELISA. RESULTS: Dyslipidemic and obese patients showed higher MCP-1 levels at month 6 from the onset of anti-TNF-α therapy (p = .05 and .01, respectively). sE-selectin positively correlated with pro-inflammatory molecules such as asymmetric dimethylarginine, sP-selectin and resistin at baseline and month 6 (p < .05). sE-selectin levels significantly reduced after 6 months of therapy (p = .0006). CONCLUSIONS: Metabolic syndrome features are associated with endothelial activation in patients with moderate-to-severe psoriasis. Adalimumab therapy led to a reduction in sE-selectin levels, supporting the beneficial effect of anti-TNF-α therapy on mechanisms associated with the development of atherosclerosis in psoriasis.