Publications by authors named "Piotr Trzonkowski"

118 Publications

Associations of TP53 codon 72 polymorphism with complications and comorbidities in patients with type 1 diabetes.

J Mol Med (Berl) 2021 Jan 25. Epub 2021 Jan 25.

Department of Medical Immunology, Faculty of Medicine, Medical University of Gdańsk, ul. Dębinki 1, 80-211, Gdańsk, Poland.

Wild-type TP53 plays an important role in the regulation of immune response and systemic inflammation. In type 1 diabetes (T1D), TP53 pathways are upregulated and an increased susceptibility to apoptosis is observed. We hypothesize that TP53 codon 72 polymorphism could be associated with complications and comorbidities in patients with T1D. We have investigated the associations of the TP53 codon 72 polymorphism with the T1D complications and comorbidities (retinopathy, nephropathy, hypertension, dyslipidemia, autoimmune thyroiditis, and celiac disease) in 350 patients. The key results of our approach are as follows: (1) In diabetic subjects, the Pro/Pro genotype is associated with an increased risk of microvascular complications, dyslipidemia, and celiac disease; (2) the Arg/Arg variant is associated with a decreased risk of autoimmune thyroiditis and celiac disease; (3) the Pro allele is associated with an increased risk of dyslipidemia, autoimmune thyroiditis, and celiac disease. Although further studies are required, our results for the first time indicate that the TP53 codon 72 polymorphism could be considered a genetic marker to predict the increased susceptibility to some T1D complications and comorbidities. KEY MESSAGES: We analyzed the TP53 codon 72 polymorphism in patients with T1D. Pro/Pro genotype is associated with an increased risk of microvascular complications, dyslipidemia, and celiac disease. The Arg/Arg variant is associated with a decreased risk of autoimmune thyroiditis and celiac disease. The Pro allele is associated with an increased risk of dyslipidemia, autoimmune thyroiditis, and celiac disease.
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http://dx.doi.org/10.1007/s00109-020-02035-1DOI Listing
January 2021

Administration of CD4CD25CD127FoxP3 Regulatory T Cells for Relapsing-Remitting Multiple Sclerosis: A Phase 1 Study.

BioDrugs 2021 Jan 5;35(1):47-60. Epub 2021 Jan 5.

Department of Medical Immunology, Medical University of Gdańsk, Debinki 7, 80-210, Gdańsk, Poland.

Background: Multiple sclerosis (MS) is an immune-mediated disease in which autoimmune T conventional (T) cells break the blood-brain barrier and destroy neurons of the central nervous system. It is hypothesized that CD4CD25CD127FoxP3 T regulatory (T) cells may inhibit this destruction through suppressive activity exerted on T cells.

Methods: We present the results of a phase 1b/2a, open-label, two-arm clinical trial in 14 patients treated with autologous T cells for relapsing-remitting MS. The patients received either expanded ex vivo T cells intravenously (intravenous [IV] group, n = 11; dose 40 × 10 T cells/kg of body weight) or freshly isolated T cells intrathecally (intrathecal [IT] group, n = 3; dose 1.0 × 10 T cells). Importantly, patients were not treated with any other disease-modifying drugs for at least 6 months before the recruitment and during the follow-up.

Results: No severe adverse events were observed. Self-assessed quality of life (EuroQol-5 Dimensions [EQ-5D] form) did not change and did not differ significantly between the groups. A total of 12 relapses were noted in five intravenously treated patients, who had from one to three attacks per year. Three out of ten participants who completed the trial in the IV group deteriorated more than 1 point on the Expanded Disability Status Scale (EDSS) during the follow-up. At the same time, no patients in the IT group experienced a relapse or such a deterioration in the EDSS. No significant differences were found in the Multiple Sclerosis Functional Composite (MSFC) scale in both the IV and IT groups. Magnetic resonance imaging (MRI) scans revealed a significantly lower change in the T2 lesion volume in the IT group compared to the IV group. The increase in the number of new T2 lesions during the follow-up was significant for the IV group only. There were no significant changes in the level of T cells or T cells in the peripheral blood throughout the follow-up or between the groups. Interestingly, T cells in all patients consisted of two different phenotypes: peripheral T cells Helios(-) (≈ 20%) and thymic T cells Helios(+) (≈ 80%). The analysis of the cytokine pattern revealed higher levels of transforming growth factor-α and proinflammatory factors MCP3, CXCL8, and IL-1RA in the IT group compared with the IV group.

Conclusions: No serious adverse events were reported in the 14 patients with MS treated with T cells in this study. The results suggest that IT administration is more promising than IV administration. Because of the low number of patients recruited, the statistical results may be underpowered and further studies are necessary to reach conclusions on efficacy and safety.

Trial Registration: EudraCT: 2014-004320-22; registered 18 November 2014.
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http://dx.doi.org/10.1007/s40259-020-00462-7DOI Listing
January 2021

Immune Status Against Hepatitis B in Patients After Allogeneic Hematopoietic Cell Transplantation-Factors Affecting Early and Long-Lasting Maintenance of Protective Anti-HBs Titers.

Front Immunol 2020 24;11:586523. Epub 2020 Nov 24.

Department of Hematology and Transplantology, Medical University of Gdansk, Gdansk, Poland.

The immunization of allogeneic hematopoietic cell transplantation (HCT) recipients against vaccine-preventable diseases is a part of posttransplantation guidelines. We conducted a prospective study to assess clinical and immunological parameters that would determine the response and long-term maintenance of protective antibody titers upon the hepatitis B virus (HBV) vaccination after HCT. The investigated variables included: vaccination of the HCT recipients and their donors prior to HCT, chronic graft versus host disease (cGVHD) and the timing of post-HCT vaccination, and B- and T-cell subtype status. Forty-two patients were immunized with three or more doses of recombinant hepatitis B surface antigen (rHBsAg) administered according to the individualized schedule of 0-1-2-6-(12) months. After vaccination, seroconversion was achieved in the whole group. The vaccines were categorized according to the antibody (Ab) titers as weak (WRs; 28.7%), good (GRs; 38%) or very good responders (VGRs; 3.3%). In multivariate logistic regression, severe cGVHD (OR= 15.5), and preceding donor immunization (OR= 0.13) were independent predictors of a weak response to vaccination. A prior belonging to the WR group impaired the durability of protection (OR= 0.17) at a median follow-up of 11.5 years. Patients with severe cGVHD showed a trend toward lower median Ab titers, although they required a higher rate of booster vaccine doses. All VGRs had CD4+ cells > 0.2 x 10/L. There was a lower mean rate of CD4+IL2+ lymphocytes in WRs. Vaccination demonstrated the immunomodulatory effect on B-cell and T-cell subsets and a Th1/Th2 cytokine profile, while shifts depended on a history of severe cGVHD and the type of vaccine responder. To conclude, vaccination of HCT donors against HBV allows a better response to vaccination in the respective HCT recipients. Double doses of rHBsAg should be considered in patients with cGVHD and in those not immunized before HCT. A dedicated intensified vaccination schedule should be administered to WRs.
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http://dx.doi.org/10.3389/fimmu.2020.586523DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7736697PMC
November 2020

Effect of serum on SmartFlare™ RNA Probes uptake and detection in cultured human cells.

Biomed J Sci Tech Res 2020 2;28(4):21788-21793. Epub 2020 Jul 2.

Department of Surgery, University of Chicago, Chicago, IL, USA.

SmartFlare™ RNA Detection Probes from Millipore is a novel technology to detect RNA in live cells based on the use of 12 nm gold nanoparticles coated with nucleotides. We proved that SmartFlares™ are internalized by human primary lymphocytes. However, fluorescence signals from target RNA detection can only be observed in the presence of Fetal Bovine Serum (FBS) in the medium, whereas it is not detectable without FBS or when medium is supplemented with human albumin. Image analysis of fluorescence generated from SmartFlare™ Uptake Control (gives constant signal regardless of contact with RNA) and RNA Specific Probes revealed further differences. In the presence of FBS, the fluorescence signal for both reagents was diffused within the cells, whereas in the absence of FBS, it was detected as single spots within the cells only when the Uptake Control was used. It is possible that FBS components are necessary for SmartFlare™ Probes to be released from cellular compartments into the cytoplasm where they can get into contact with target RNA. The exact mechanism of this phenomena should be further determined. However, for the first time, we present here that FBS in the cell culture medium is essential for RNA detection by SmartFlare™ technology in human lymphocytes.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7447198PMC
July 2020

Antigen-reactive regulatory T cells can be expanded in vitro with monocytes and anti-CD28 and anti-CD154 antibodies.

Cytotherapy 2020 11 7;22(11):629-641. Epub 2020 Aug 7.

Department of Medical Immunology, Medical University of Gdansk, Gdańsk, Poland; Poltreg S.A., Gdańsk, Poland. Electronic address:

Background: In recent years, therapies with CD4CD25FoxP3 regulatory T cells (Tregs) have been successfully tested in many clinical trials. The important issue regarding the use of this treatment in autoimmune conditions remains the specificity toward particular antigen, as because of epitope spread, there are usually multiple causative autoantigens to be regulated in such conditions.

Methods: Here we show a method of generation of Tregs enriched with antigen-reactive clones that potentially covers the majority of such autoantigens. In our research, Tregs were expanded with anti-CD28 and anti-CD154 antibodies and autologous monocytes and loaded with a model peptide, such as whole insulin or insulin β chain peptide 9-23. The cells were then sorted into cells recognizing the presented antigen. The reactivity was verified with functional assays in which Tregs suppressed proliferation or interferon gamma production of autologous effector T cells (polyclonal and antigen-specific) used as responders challenged with the model peptide. Finally, we analyzed clonotype distribution and TRAV gene usage in the specific Tregs.

Results: Altogether, the applied technique had a good yield and allowed us to obtain a Treg product enriched with a specific subset, as confirmed in the functional tests. The product consisted of many clones; nevertheless, the content of these clones was different from that found in polyclonal or unspecific Tregs.

Conclusions: The presented technique might be used to generate populations of Tregs enriched with cells reactive to any given peptide, which can be used as a cellular therapy medicinal product in antigen-targeted therapies.
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http://dx.doi.org/10.1016/j.jcyt.2020.07.001DOI Listing
November 2020

Podocytes-The Most Vulnerable Renal Cells in Preeclampsia.

Int J Mol Sci 2020 Jul 17;21(14). Epub 2020 Jul 17.

Department of Obstetrics and Gynecology, Pomeranian Medical University, 70-111 Szczecin, Poland.

Preeclampsia (PE) is a disorder that affects 3-5% of normal pregnancies. It was believed for a long time that the kidney, similarly to all vessels in the whole system, only sustained endothelial damage. The current knowledge gives rise to a presumption that the main role in the development of proteinuria is played by damage to the podocytes and their slit diaphragm. The podocyte damage mechanism in preeclampsia is connected to free VEGF and nitric oxide (NO) deficiency, and an increased concentration of endothelin-1 and oxidative stress. From national cohort studies, we know that women who had preeclampsia in at least one pregnancy carried five times the risk of developing end-stage renal disease (ESRD) when compared to women with physiological pregnancies. The focal segmental glomerulosclerosis (FSGS) is the dominant histopathological lesion in women with a history of PE. The kidney's podocytes are not subject to replacement or proliferation. Podocyte depletion exceeding 20% resulted in FSGS, which is a reason for the later development of ESRD. In this review, we present the mechanism of kidney (especially podocytes) injury in preeclampsia. We try to explain how this damage affects further changes in the morphology and function of the kidneys after pregnancy.
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http://dx.doi.org/10.3390/ijms21145051DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7403979PMC
July 2020

Comparisons of Dipstick Test, Urine Protein-to-Creatine Ratio, and Total Protein Measurement for the Diagnosis of Preeclampsia.

Int J Environ Res Public Health 2020 06 12;17(12). Epub 2020 Jun 12.

Cancer Immunology Group, International Centre for Cancer Vaccine Science, University of Gdansk, 80-214 Gdańsk, Poland.

Preeclampsia affects 2-5% of pregnant women and is one of the leading causes of maternal and perinatal morbidity and mortality. We aimed to extensively evaluate proteinuria in women with preeclampsia and to determine the analytical sensitivity and specificity of and the cutoff values for urine protein-to-creatinine ratio (UPCR) and total protein in 24 h urine samples. This study included 88 women. We used the urine dipstick test, UPCR, and total protein measurement in a 24 h urine sample. The patients were divided in gestational hypertension (GH, = 44) and preeclampsia (PE, = 44) groups. In the GH group, 25% (11/44) of the patients presented incidentally positive results. UPCR and total protein in 24 h urine specimens were increased in the GH group compared to the PE group. Receiver operating characteristic analysis showed a UPCR cutoff of 30 mg/mmol as significant for preeclampsia, while the sensitivity and specificity were 89% (95% CI, 75-97) and 100% (95% CI, 87-100), respectively. In the 24 h urine protein test, sensitivity and specificity were 80% (95% CI, 61-92) and 100% (95% CI, 88-100), respectively, for the cutoff value of 0.26 g/24 h. In comparison to the other commonly used tests here considered, UPCR determination is a reliable, relatively faster, and equally accurate method for the quantitation of proteinuria, correlates well with 24 h urine protein estimations, and could be used as an alternative to the 24 h proteinuria test for the diagnosis of preeclampsia.
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http://dx.doi.org/10.3390/ijerph17124195DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7344421PMC
June 2020

COVID-19 - Toward a comprehensive understanding of the disease.

Cardiol J 2020 7;27(2):99-114. Epub 2020 May 7.

1st Department of Cardiology, University Catheterization Laboratories, Medical University of Gdansk, Poland.

The evidence on the pathophysiology of the novel coronavirus SARS-CoV-2 infection is rapidly growing. Understanding why some patients suffering from COVID-19 are getting so sick, while others are not, has become an informal imperative for researchers and clinicians around the globe. The answer to this question would allow rationalizing the fear surrounding this pandemic. Understanding of the pathophysiology of COVID-19 relies on an understanding of interplaying mechanisms, including SARS-CoV-2 virulence, human immune response, and complex inflammatory reactions with coagulation playing a major role. An interplay with bacterial co-infections, as well as the vascular system and microcirculation affected throughout the body should also be examined. More importantly, a compre-hensive understanding of pathological mechanisms of COVID-19 will increase the efficacy of therapy and decrease mortality. Herewith, presented is the current state of knowledge on COVID-19: beginning from the virus, its transmission, and mechanisms of entry into the human body, through the pathological effects on the cellular level, up to immunological reaction, systemic and organ presentation. Last but not least, currently available and possible future therapeutic and diagnostic options are briefly commented on.
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http://dx.doi.org/10.5603/CJ.a2020.0065DOI Listing
June 2020

GPA33: A Marker to Identify Stable Human Regulatory T Cells.

J Immunol 2020 06 4;204(12):3139-3148. Epub 2020 May 4.

Department of Hematopoiesis, Sanquin Research and Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam, 1066 CX Amsterdam, the Netherlands;

FOXP3-expressing regulatory T (Treg) cells safeguard immunological tolerance. Treg cells can be generated during thymic development (called thymic Treg [tTreg] cells) or derived from mature conventional CD4 T cells that underwent TGF-β-mediated conversion in the periphery (called peripheral Treg [pTreg] cells). Murine studies have shown that tTreg cells exhibit strong lineage fidelity, whereas pTreg cells can revert into conventional CD4 T cells. Their stronger lineage commitment makes tTreg cells the safest cells to use in adoptive cell therapy, increasingly used to treat autoimmune and inflammatory disorders. Markers to distinguish human tTreg cells from pTreg cells have, however, not been found. Based on combined proteomic and transcriptomic approaches, we report that the Ig superfamily protein GPA33 is expressed on a subset of human Treg cells. GPA33 is acquired late during tTreg cell development but is not expressed on TGF-β-induced Treg cells. GPA33 identifies Treg cells in human blood that lack the ability to produce effector cytokines (IL-2, IFN-γ, IL-17), regardless of differentiation stage. GPA33 Treg cells universally express the transcription factor Helios that preferentially marks tTreg cells and can robustly and stably be expanded in vitro even without rapamycin. Expanded GPA33 Treg cells are suppressive, unable to produce proinflammatory cytokines, and exhibit the epigenetic modifications of the FOXP3 gene enhancer CNS2, necessary for indelible expression of this critical transcription factor. Our findings thus suggest that GPA33 identifies human tTreg cells and provide a strategy to isolate such cells for safer and more efficacious adoptive cell therapy.
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http://dx.doi.org/10.4049/jimmunol.1901250DOI Listing
June 2020

Immune Algorithm Optimization for Organ Transplantation in Poland.

Transplant Proc 2020 Sep 9;52(7):2026-2032. Epub 2020 Apr 9.

Department of Medical Immunology, Medical University of Gdańsk, Gdańsk, Poland.

The lack of a uniform method for determining unacceptable HLA mismatches (UAMs) for organ transplantation worldwide has resulted in many different algorithms for donor-recipient matching. Here we present our proposal for changes to the current algorithm for immune evaluation of potential kidney recipients in Poland based on the experience of various transplantation centers. The most important finding of this article is an algorithm that stratifies the pretransplant immunologic risk based on strict laboratory criteria, enabling harmonization between transplant centers in Poland. This is because of a step-by-step algorithm for alloantibody assessment using solid-phase assays (SPA) and clearly defined technical issues, as well as cutoffs for reporting UAMs. Our novel approach focuses on a laboratory testing extension in the scope of HLA typing; detection and characterization of alloantibodies before transplantation; desensitization; and post-transplant monitoring. The proposed changes will allow for the assessment of clinically relevant anti-HLA antibodies with complement binding properties; the determination of UAMs in the potential donor; the calculation of virtual panel reactive antibodies (vPRA); the calculation of the recipient's immunologic rejection risk stratification; the assessment of the donor-recipient virtual cross-match (vXM); and the determination of the final recipient's selection for the biological cross-match testing. Collectively, the optimized algorithm permit for UAM verification is based on laboratory proofed data and will firmly improve organ allocation and transplant outcomes in Poland. We hope that this novel approach also improves the individual patient's risk stratification and future personalized treatment.
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http://dx.doi.org/10.1016/j.transproceed.2020.02.082DOI Listing
September 2020

Proinsulin-specific T regulatory cells may control immune responses in type 1 diabetes: implications for adoptive therapy.

BMJ Open Diabetes Res Care 2020 02;8(1)

Department of Medical Immunology, Medical University of Gdańsk, Gdańsk, Poland

Objective: Here we looked for possible mechanisms regulating the progression of type 1 diabetes mellitus (T1DM). In this disease, autoaggressive T cells (T conventional cells, Tconvs) not properly controlled by T regulatory cells (Tregs) destroy pancreatic islets.

Research Design And Methods: We compared the T-cell compartment of patients with newly diagnosed T1DM (NDT1DM) with long-duration T1DM (LDT1DM) ones. The third group consisted of patients with LDT1DM treated previously with polyclonal Tregs (LDT1DM with Tregs). We have also looked if the differences might be dependent on the antigen specificity of Tregs expanded for clinical use and autologous sentinel Tconvs.

Results: Patients with LDT1DM were characterized by T-cell immunosenescence-like changes and expansion of similar vβ/T-cell receptor (TCR) clones in Tconvs and Tregs. The treatment with Tregs was associated with some inhibition of these effects. Patients with LDT1DM possessed an increased percentage of various proinsulin-specific T cells but not GAD65-specific ones. The percentages of all antigen-specific subsets were higher in the expansion cultures than in the peripheral blood. The proliferation was more intense in proinsulin-specific Tconvs than in specific Tregs but the levels of some proinsulin-specific Tregs were exceptionally high at baseline and remained higher in the expanded clinical product than the levels of respective Tconvs in sentinel cultures.

Conclusions: T1DM is associated with immunosenescence-like changes and reduced diversity of T-cell clones. Preferential expansion of the same TCR families in both Tconvs and Tregs suggests a common trigger/autoantigen responsible. Interestingly, the therapy with polyclonal Tregs was associated with some inhibition of these effects. Proinsulin-specific Tregs appeared to be dominant in the immune responses in patients with T1DM and probably associated with better control over respective autoimmune Tconvs.

Trial Registration Number: EudraCT 2014-004319-35.
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http://dx.doi.org/10.1136/bmjdrc-2019-000873DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7206972PMC
February 2020

Immunosuppressive properties of amino acid and peptide derivatives of mycophenolic acid.

Eur J Med Chem 2020 Mar 24;189:112091. Epub 2020 Jan 24.

Department of Organic Chemistry, Gdansk University of Technology, G. Narutowicza 11/12, 80-233, Gdansk, Poland.

Mycophenolic acid (MPA) was coupled with amino acids and biologically active peptides including derivatives of tuftsin to modify its immunosuppressive properties. Both amino acid unit in the case of simple MPA amides and modifications within peptide moiety of MPA - tuftsin conjugates influenced the observed activity. Antiproliferative potential of the obtained conjugates was investigated in vitro and MPA amides with threonine methyl ester and conjugate of MPA with retro-tuftisin occurred to be more selective against PBMC in comparison to parent MPA. Both amino acid and peptide derivatives of MPA acted as inosine-5'-monophosphate dehydrogenaze (IMPDH) inhibitors.
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http://dx.doi.org/10.1016/j.ejmech.2020.112091DOI Listing
March 2020

Changing microRNA Expression during Three-Month Wasp Venom Immunotherapy.

Immunol Invest 2019 Nov 24;48(8):835-843. Epub 2019 May 24.

Department of Allergology, Medical University of Gdansk , Gdansk , Poland.

MicroRNAs are small non-coding molecules playing a significant regulatory role in several allergic diseases. However their role in tolerance induction remains unclear. The aim of this study was to determine the expression of selected microRNAs during the first three months of wasp venom immunotherapy (VIT). 5 adult patients with a history of severe systemic reactions after stinging by wasps and confirmed sensitization were included. Venous blood samples were collected before VIT, 24 hours after completing its initial phase and after 3 months of the maintenance therapy. A control group was comprised of 5 healthy individuals with no history of allergy. In the blood samples expression of 96 microRNAs was determined with the use of microfluidic cards. In a statistical analysis the expression was compared between the study groups as well as between the pre- and post-VIT samples. Significant differences were found between the patients with wasp venom allergy and the healthy controls in the expression of miR-601 and miR-1201 upregulated in allergic patients at every time point ( = 0.04; = 0.015, respectively). During VIT profile of microRNA was changing with lower expression of 6 microRNAs (including miR-182, miR-342, miR-375) and higher of 11 microRNAs (including let-7d, miR-34b, miR-143). To conclude, VIT has led to some changes in the expression of microRNA associated with Th2-type inflammation and tolerance induction.
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http://dx.doi.org/10.1080/08820139.2019.1617303DOI Listing
November 2019

Regulatory T cells: the future of autoimmune disease treatment.

Expert Rev Clin Immunol 2019 07 27;15(7):777-789. Epub 2019 May 27.

a Department of Medical Immunology , Laboratory of Experimental Immunology, Medical University of Gdańsk , Debinki , Poland.

: CD4 + T regulatory cells (Tregs) have been described as the most potent immunosuppressive cells in the human body. They have been found to control autoimmunity, and clinical attempts have been made to apply them to treat autoimmune diseases. Some specific pathways utilized by Tregs in the regulation of immune response or Tregs directly as cellular products are tested in the clinic. : Here, we present recent advances in the research on the biology and clinical applications of Tregs in the treatment of autoimmune diseases. : Regulatory T cells seem to be a promising tool for the treatment of autoimmune diseases. The development of both cell-based therapies and modern pharmacotherapies which affect Tregs may strongly improve the treatment of autoimmune disorders. Growing knowledge about Treg biology together with the latest biotechnology tools may give an opportunity for personalized therapies in these conditions.
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http://dx.doi.org/10.1080/1744666X.2019.1620602DOI Listing
July 2019

Therapy with CD4CD25 T regulatory cells - should we be afraid of cancer?

Contemp Oncol (Pozn) 2019 5;23(1):1-6. Epub 2019 Apr 5.

Department of Medical Immunology, Medical University of Gdansk, Gdansk, Poland.

This review focuses on the role of regulatory T cells (Tregs) in the process of carcinogenesis. The controversy of this issue arose due to the increasing therapeutic use of Tregs in humans (inter alia, in the treatment of autoimmune diseases). It is mainly due to potential dangers related to immunosuppressive activity of these cells, especially regarding cancer. The natural function of regulatory T cells (which is the suppression of excessive activity of the immune system) is purportedly linked to an increased risk of cancer initiation. This work brings together and summarizes the most important reports of researchers dealing with this problem and attempts to explain doubts and fears related to Tregs and their uncertain connection with cancer initiation and progression. It is clearly shown that regulatory T cells are associated with acceleration of existing tumors (they are attracted by microenvironments created by cancer cells) but cannot initiate them on their own.
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http://dx.doi.org/10.5114/wo.2019.84110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6500397PMC
April 2019

Ways Forward for Tolerance-Inducing Cellular Therapies- an AFACTT Perspective.

Front Immunol 2019 22;10:181. Epub 2019 Feb 22.

Division of Immunology, Germans Trias i Pujol University Hospital, LCMN, IGTP, Badalona, Spain.

Clinical studies with cellular therapies using tolerance-inducing cells, such as tolerogenic antigen-presenting cells (tolAPC) and regulatory T cells (Treg) for the prevention of transplant rejection and the treatment of autoimmune diseases have been expanding the last decade. In this perspective, we will summarize the current perspectives of the clinical application of both tolAPC and Treg, and will address future directions and the importance of immunomonitoring in clinical studies that will result in progress in the field.
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http://dx.doi.org/10.3389/fimmu.2019.00181DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6395407PMC
January 2020

Synthesis and antiproliferative activity of new mycophenolic acid conjugates with adenosine derivatives.

J Asian Nat Prod Res 2019 Feb 2;21(2):178-185. Epub 2018 Apr 2.

a Department of Organic Chemistry , Gdansk University of Technology , Gdansk PL 80-233 , Poland.

New conjugates of mycophenolic acid (MPA) and adenosine derivatives were synthesized and assessed as potential immunosuppressants on Jurkat cell line and peripheral blood mononuclear cells (PBMC) from healthy donors. As compared to MPA, all compounds were found to be more active against Jurkat cell line. The antiproliferative activities were compared with MPA and adenosine, in both 2',3'-O-isopropylidene protected and free hydroxyl groups possessing forms. The obtained results were also discussed in terms of selectivity index, defined as SI = IC/EC.
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http://dx.doi.org/10.1080/10286020.2018.1451521DOI Listing
February 2019

Cell banking for regulatory T cell-based therapy: strategies to overcome the impact of cryopreservation on the Treg viability and phenotype.

Oncotarget 2018 Feb 3;9(11):9728-9740. Epub 2018 Jan 3.

Department of Surgery, University of Chicago, Chicago, IL, USA.

The first clinical trials with adoptive Treg therapy have shown safety and potential efficacy. Feasibility of such therapy could be improved if cells are cryopreserved and stored until optimal timing for infusion. Herein, we report the evaluation of two cell-banking strategies for Treg therapy: 1) cryopreservation of CD4 cells for subsequent Treg isolation/expansion and 2) cryopreservation of expanded Tregs (CD4CD25CD127 cells). First, we checked how cryopreservation affects cell viability and Treg markers expression. Then, we performed Treg isolation/expansion with the final products release testing. We observed substantial decrease in cell number recovery after thawing and overnight culture. This observation might be explained by the high percentage of necrotic and apoptotic cells found just after thawing. Furthermore, we noticed fluctuations in percentage of CD4CD25CD127 and CD4FoxP3 cells obtained from cryopreserved CD4 as well as Treg cells. However, after re-stimulation Tregs expanded well, presented a stable phenotype and fulfilled the release criteria at the end of expansions. Cryopreservation of CD4 cells for subsequent Treg isolation/expansion and cryopreservation of expanded Tregs with re-stimulation and expansion after thawing, are promising solutions to overcome detrimental effects of cryopreservation. Both of these cell-banking strategies for Treg therapy can be applied when designing new clinical trials.
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http://dx.doi.org/10.18632/oncotarget.23887DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5839397PMC
February 2018

Mast cell derived carboxypeptidase A3 is decreased among patients with advanced coronary artery disease.

Cardiol J 2019 7;26(6):680-686. Epub 2018 Mar 7.

Laboratory of Immunoregulation and Cellular Therapies, Department of Family Medicine, Medical University of Gdansk, Gdansk, Poland.

Background: Coronary artery disease (CAD) affects milions of people and can result in myocardial infarction (MI). Previously, mast cells (MC) have been extensively investigated in the context of hypersensitivity, however as regulators of the local inflammatory response they can potentially contribute to CAD and/or its progression. The aim of the study was to assess if serum concentration of MC proteases: carboxypeptidase A3, cathepsin G and chymase 1 is associated with the extension of CAD and MI.

Methods: The 44 patients with angiographically confirmed CAD (23 subjects with non-ST-segment elevation MI [NSTEMI] and 21 with stable CAD) were analyzed. Clinical data were obtained as well serum concentrations of carboxypeptidase A3, cathepsin G and chymase 1 were also measured.

Results: Patients with single vessel CAD had higher serum concentration of carboxypeptidase than those with more advanced CAD (3838.6 ± 1083.1 pg/mL vs. 2715.6 ± 442.5 pg/mL; p = 0.02). There were no significant differences in levels of any protease between patients with stable CAD and those with NSTEMI. Patients with hypertension had ≈2-fold lower serum levels of cathepsin G than normotensive individuals (4.6 ± 0.9 pg/mL vs. 9.4 ± 5.8 pg/mL; p = 0.001). Cathepsin G levels were also decreased in sera of the current smokers as compared with non-smokers (3.1 ± 1.2 ng/mL vs. 5.8 ± 1.2 ng/mL, p = 0.02).

Conclusions: Decreased serum level of carboxypeptidase is a hallmark of more advanced CAD. Lower serum levels of carboxypeptidase A3 and catepsin G are associated with risk factors of blood vessel damage suggesting a protective role of these enzymes in CAD.
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http://dx.doi.org/10.5603/CJ.a2018.0018DOI Listing
August 2020

The role of regulatory T cells and genes involved in their differentiation in pathogenesis of selected inflammatory and neoplastic skin diseases. Part II: The Treg role in skin diseases pathogenesis.

Postepy Dermatol Alergol 2017 Oct 31;34(5):405-417. Epub 2017 Oct 31.

Department of Dermatology, Venereology and Allergology, Medical University of Gdansk, Gdansk, Poland.

Regulatory FOXP3+ T cells (Tregs) constitute 5% to 10% of T cells in the normal human skin. They play an important role in the induction and maintenance of immunological tolerance. The suppressive effects of these cells are exerted by various mechanisms including the direct cytotoxic effect, anti-inflammatory cytokines, metabolic disruption, and modulation of the dendritic cells function. The deficiency of Treg cells number or function are one of the basic elements of the pathogenesis of many skin diseases, such as psoriasis, atopic dermatitis, bacterial and viral infections. They also play a role in the pathogenesis of T cell lymphomas of the skin (cutaneous T cell lymphomas - CTCL), skin tumors and mastocytosis. Here, in the second part of the cycle, we describe dysfunctions of Tregs in selected skin diseases.
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http://dx.doi.org/10.5114/ada.2017.71105DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5835974PMC
October 2017

Peripheral regulatory T cells and anti-inflammatory cytokines in children with juvenile idiopathic arthritis.

Acta Biochim Pol 2018 1;65(1):119-123. Epub 2018 Mar 1.

Department of Pediatrics, Pediatric Gastroenterology, Hepatology and Nutrition, Medical University of Gdańsk, Gdańsk, Poland.

Background: Juvenile idiopathic arthritis (JIA) is a chronic, heterogenous inflammatory disease of unclear pathogenesis. JIA is hypothesized to be linked to a defective immune regulation. Anti-inflammatory cytokines belong to the best known regulatory factors. T-regulatory cells are a crucial cellular component of immune tolerance. One of their functions is synthesis of interleukin 10 (IL-10) and transforming growth factor beta1 (TGF-β1). The aim of this study was to determine the proportion of T-regulatory cells (CD4CD25FOXP3) in peripheral blood, and serum levels of TGF-β1 and IL-10 in patients with JIA.

Methods: The study included 25 patients with newly diagnosed JIA: oligoarthritis (n=17) and polyarthritis (n=8). The control group was comprised of 17 healthy children. CD4CD25FOXP3 T cells in peripheral blood were quantified by means of three-color flow cytometry. Serum concentrations of TGF-β1 and IL-10 were estimated with ELISA.

Results: The proportion of peripheral CD4CD25FOXP3 cells in patients with JIA was significantly higher than in the controls (p=0.04). The two groups did not differ significantly in terms of their TGF-β1 and IL-10 concentrations.

Conclusions: At the time of diagnosis, children with JIA presented with an elevated proportion of T-regulatory cells (CD4CD25FOXP3) in peripheral blood. Anti-inflammatory cytokines, IL-10 and TGF-β1, are not upregulated in the serum of patients with JIA, and therefore should not be considered as biomarkers of this condition.
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http://dx.doi.org/10.18388/abp.2017_2308DOI Listing
October 2018

The role of regulatory T cells and genes involved in their differentiation in pathogenesis of selected inflammatory and neoplastic skin diseases. Part III: Polymorphisms of genes involved in Tregs' activation and function.

Postepy Dermatol Alergol 2017 Dec 31;34(6):517-525. Epub 2017 Dec 31.

Department of Dermatology, Venereology and Allergology, Medical University of Gdansk, Gdansk, Poland.

Regulatory T cells (Tregs) represent a cell type that promotes immune tolerance to autologous components and maintains immune system homeostasis. The abnormal function of Tregs is relevant to the pathogenesis of several skin diseases like psoriasis, atopic dermatitis, systemic lupus erythematosus, cutaneous T-cell lymphomas, and skin cancer and is also important in rheumatoid arthritis, diabetes and other autoimmune diseases. In this review, we will summarize the role of mutations and/or polymorphisms of genes involved in Tregs development, and functions in the pathogenesis of selected skin diseases.
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http://dx.doi.org/10.5114/pdia.2017.67053DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5799752PMC
December 2017

Minimum Information about T Regulatory Cells: A Step toward Reproducibility and Standardization.

Front Immunol 2017 15;8:1844. Epub 2018 Jan 15.

Department of Clinical Immunology and Transplantology, Medical University of Gdańsk, Gdańsk, Poland.

Cellular therapies with CD4+ T regulatory cells (Tregs) hold promise of efficacious treatment for the variety of autoimmune and allergic diseases as well as posttransplant complications. Nevertheless, current manufacturing of Tregs as a cellular medicinal product varies between different laboratories, which in turn hampers precise comparisons of the results between the studies performed. While the number of clinical trials testing Tregs is already substantial, it seems to be crucial to provide some standardized characteristics of Treg products in order to minimize the problem. We have previously developed reporting guidelines called minimum information about tolerogenic antigen-presenting cells, which allows the comparison between different preparations of tolerance-inducing antigen-presenting cells. Having this experience, here we describe another minimum information about Tregs (MITREG). It is important to note that MITREG does not dictate how investigators should generate or characterize Tregs, but it does require investigators to report their Treg data in a consistent and transparent manner. We hope this will, therefore, be a useful tool facilitating standardized reporting on the manufacturing of Tregs, either for research purposes or for clinical application. This way MITREG might also be an important step toward more standardized and reproducible testing of the Tregs preparations in clinical applications.
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http://dx.doi.org/10.3389/fimmu.2017.01844DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5775516PMC
January 2018

Transcriptional profile of in vitro expanded human epidermal progenitor cells for the treatment of non-healing wounds.

J Dermatol Sci 2018 Mar 13;89(3):272-281. Epub 2017 Dec 13.

Department of Clinical Immunology and Transplantology, Medical University of Gdansk, Poland. Electronic address:

Background: Epidermal progenitor cells (EPCs) have been under extensive investigation due to their increasing potential of application in medicine and biotechnology. Cultured human EPCs are used in the treatment of chronic wounds and have recently became a target for gene therapy and toxicological studies. One of the challenges in EPCs culture is to provide a high number of undifferentiated, progenitor cells displaying high viability and significant biological activity.

Objectives: The goal of this study was to characterize the in vitro cultured progenitor cells and to assess whether the cells with the progenitor phenotype are able to enhance wound healing. Additionally, we aimed to establish the complete procedure of the culture, analysis and clinical application of epidermal progenitor cells.

Methods: In this study we present a method of cell isolation and culture followed by a technique of transplantation of the cultured cells onto the wound bed. The applied isolation technique involves two enzymatic steps (dispase, trypsin) and it is characterized by a high yield of cells. The obtained cells were cultured in vitro up to the second passage in serum-free and xeno-free keratinocytes-dedicated medium. Key stem cell markers were determined with means of flow cytometry and quantitative real-time PCR.

Results: The in vitro expanded cells displayed high proliferative activity without features of neither apoptosis nor necrosis. The flow cytometry and transcriptomic analyses showed enhanced expression of stem cell markers (i.e. proteins: ΔNp63, CD29, CD49f and BNC1, CDKN1A transcripts) in the expanded cells. In the presented compassionate use study, cultured autologous cells from an oncological patient were suspended in fibrin sealant and transplanted directly to a non-healing wound, resulting in wound closure within 2 months.

Conclusion: The cells cultured in serum-free media display epidermal stem cells features and a potential to stimulate wound healing. This promising procedure of isolation, culture and application warrants further clinical trials in the treatment of chronic wounds.
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http://dx.doi.org/10.1016/j.jdermsci.2017.12.003DOI Listing
March 2018

Author Correction: Mild hypothermia provides Treg stability.

Sci Rep 2017 12 15;7(1):17940. Epub 2017 Dec 15.

Department of Clinical Immunology and Transplantology, Medical University of Gdańsk, ul. Dębinki 7, 80-210, Gdańsk, Poland.

A correction to this article has been published and is linked from the HTML version of this paper. The error has been fixed in the paper.
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http://dx.doi.org/10.1038/s41598-017-17186-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5732292PMC
December 2017

Impact of donor and recipient human cytomegalovirus status on kidney transplantation.

Int Immunol 2017 12;29(12):541-549

Department of Clinical Immunology and Transplantology, Medical University of Gdansk, Gdansk, Poland.

Human cytomegalovirus (HCMV) is considered to be a major pathogen that affects the outcome of solid organ transplantation (TX). Both recipient and donor may be HCMV positive, therefore HCMV re-infection is possible after TX. However, little is known how cytomegalovirus (CMV) transmitted from an infected donor to an infected recipient modulates the recipient's already suppressed immunity, and what the clinical consequences are. To investigate these issues, 52 kidney recipients were followed up for 2 years after TX. T, B and natural killer (NK) lymphocytes, naive and memory T subsets, CD28 expression, relative telomere length, CMV-specific lymphocytes and serum cytokines were measured several times post-TX. Patients were monitored for signs of CMV viremia and other infections. The most important observation was that CMV-specific lymphocytes expand vastly in HCMV-infected recipients who received kidneys from infected donors, in comparison with uninfected donors. Despite this, a higher rate of HCMV viremia was found. Immune deterioration was confirmed by an increased number of CD28-negative T lymphocytes, inverted CD4/CD8 index and shortened telomeres. This was superior in HCMV-infected recipients transplanted from infected donors, when compared with uninfected. In conclusion, CMV alters the immune system in kidney transplant recipients and promotes immune exhaustion.
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http://dx.doi.org/10.1093/intimm/dxx062DOI Listing
December 2017

Association of cytokine gene polymorphisms with the complications of allogeneic haematopoietic stem cell transplantation.

Hum Immunol 2017 Nov 4;78(11-12):672-683. Epub 2017 Oct 4.

Department of Clinical Immunology and Transplantology, Medical University of Gdansk, Debinki 7 Street, 80-211 Gdansk, Poland. Electronic address:

The purpose of our study was to confirm the prevalence of the association between single nucleotide polymorphisms present in genes encoding cytokines and the complications occurring after haematopoietic stem cell transplantation (HSCT). 108 recipients and 81 donors were typed for TNF-α (-308), TGF-β1 (codon 10, 25), IL-10 (-1082, -819, -592), IL-6 (-174) and INF-γ (+874). Our studies have shown a tendency toward association between the occurrence of acute form of graft versus host disease (aGVHD) and IL-6 genotype. Homozygote C/C was less likely to develop aGVHD (p=0,09). Genotype GCC/ATA in IL-10 recipient gene alone had protective effect against the occurrence of aGVHD (p=0,01). Furthermore, GCC/ATA protected the host against developing the disease in the clinically relevant grades (II-IV) (p=0,03). In addition, the recipient's T/T G/G genotype (TGF-β1) predisposed to the development of both acute (p=0,06 - trend) and chronic (p=0,04) GVHD and also severe aGVHD (p=0,004). We also observed a statistically significant association between the genotype of recipient and the risk of infection - the protective function of the G/C IL-6 in the bloodstream infections (p=0,001). Our results suggest that IL-6, IL-10 and TGF-β1 genotypes of recipient are the most associated with the risk of complications after HSCT.
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http://dx.doi.org/10.1016/j.humimm.2017.09.005DOI Listing
November 2017

The role of regulatory T cells and genes involved in their differentiation in pathogenesis of selected inflammatory and neoplastic skin diseases. Part I: Treg properties and functions.

Postepy Dermatol Alergol 2017 Aug 31;34(4):285-294. Epub 2017 Jul 31.

Department of Dermatology, Venereology and Allergology, Medical University of Gdansk, Gdansk, Poland.

Regulatory T cells (Treg) can be divided into two types: the natural cells (tTreg), which arise in the thymus, and the induced cells (iTreg), which are produced in peripheral tissues during immune response. The most recently published studies indicate that the supervisory functions of these cells are weakened in the pathogenesis of autoimmune and neoplastic diseases of the skin. This may be a result of the domination of other immune cells in the skin, such as Th1/Th17/Th22 and Tc1 type in psoriasis and Th2 in atopic dermatitis. The excessive activity of Treg cells can lead to immunosuppression and decrease in the number of Th1 cells, which promote the development and progression of skin cancers. In the case of cutaneous T-cell lymphomas, there are suggestions that tumor progression is associated with the acquisition of the suppressor phenotype of malignant cells. There is genetic background of Treg dysfunction in skin disorders. This article describes the types and functions of Treg cells.
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http://dx.doi.org/10.5114/ada.2017.69305DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5560174PMC
August 2017

Mild hypothermia provides Treg stability.

Sci Rep 2017 09 20;7(1):11915. Epub 2017 Sep 20.

Department of Clinical Immunology and Transplantology, Medical University of Gdańsk, ul. Dębinki 7, 80-210, Gdańsk, Poland.

Regulatory T cells (Tregs) play crucial role in maintenance of peripheral tolerance. Recent clinical trials confirmed safety and efficacy of Treg treatment of deleterious immune responses. However, Tregs lose their characteristic phenotype and suppressive potential during expansion ex vivo. Therefore, multiple research teams have been studding Treg biology in aim to improve their stability in vitro. In the current paper, we demonstrate that mild hypothermia of 33 °C induces robust proliferation of Tregs, preserves expression of FoxP3, CD25 and Helios, and prevents TSDR methylation during culture in vitro. Tregs expanded at 33 °C have stronger immunosuppressive potential and remarkably anti-inflammatory phenotype demonstrated by the whole transcriptome sequencing. These observations shed new light on impact of temperature on regulation of immune response. We show that just a simple change in temperature can preserve Treg stability, function and accelerate their proliferation, responding to unanswered question- how to preserve Treg stability in vitro.
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http://dx.doi.org/10.1038/s41598-017-10151-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5607276PMC
September 2017