Resident Memory T Cells

Until recently, T cells were thought to remain in circulation until recruitment of the inflammation and only a small number of T cells remained in the peripheral tissues without inflammation. However, studies have found that a group of T cells settled in the tissues and remained there for a long time. Those cells are named as tissue-resident memory T cells (TRM). TRM cells are transcriptionally, phenotypically, and functionally distinct from other T cells, which recirculate between blood, secondary lymphoid organs, and non-lymphoid tissues. They undergo a distinct proliferation that discriminates them from circulating T cells and their main cell surface markers are CD69, CD103, and CD49a. Upon exposure to the same or similar diseases, TRM cells provide a first line of adaptive cellular defense against infection in peripheral non-lymphoid tissues, such as skin, lungs, digestive, and urogenital tracts. This approach forms the basis of a novel vaccination strategy called “prime and pull”, which ensures long-term local immunity. On the other hand, abnormal activated and malignant TRM may contribute to numerous human inflammatory diseases such as psoriasis and vitiligo. Here in this chapter, we aimed to emphasize TRM cell location, migration, phenotypic structure, maintenance, and diseases associated with TRM cells

Optimization and screening of solid lipid nanoparticle production for gene delivery by factorial design and response surface methodology

Aim: A successful gene therapy requires a delivery system for overcoming various biological barriers. For this, we adapted the factorial design and response surface methodology to the cationic solid lipid nanoparticle production process. Methods: Screening and optimization of formulations were carried out with factorial design with 3 factors and 3 levels using Box-Behnken Design. Then, solid lipid nanoparticles were physicochemically characterized. Furthermore, optimal SLN formulation is examined in terms of complex formation with plasmid DNA, its protection potential against nucleases, cytotoxicity profile, and storage stability. Results: Response-surface analyses demonstrated that the selected quadratic model holds significance for particle size and zeta potential. The interaction of independent variables was statistically determined. Optimization and prediction were performed using obtained second-order polynomial equations. Optimal formulation and complexes were found to be nanosized, positively charged and their polydispersity-index values below 0.3 as an indicator of being monodispersed.  Cytotoxicity of the optimal formulation is compatible for further studies and no significant increase was observed in particle size until day 21 and until day 60 for polydispersity-index. Conclusion: Optimal formulation provides a good basis as a gene delivery system was produced with developed systematic. Briefly, this methodology could be used to obtain SLNs with desired conditions

Pagophagia-Induced Hyponatremia: An Unusual Case

Hyponatremia occurs when the serum sodium level is below 135 mmol/L. The symptoms include nausea, vomiting, confusion, headache, cardiorespiratory symptoms, profound somnolence or coma, and seizures are observed. Iron deficiency anemia can also cause pagophagia, a Pica subspecies. Although it has been emphasized that electrolyte disturbance may develop due to pagophagia, only a dearth of cases was reported. A 59-year-old male patient was brought to the emergency department with complaints of incoherent speech that started at night, disorientated movements (such as fluttering and climbing), insomnia, restlessness, and confusion. In 2017, he experienced hyponatremia due to pagophagia and a salt-free diet. At the index episode of hyponatremia, he experienced confusion, drowsiness, and sleepiness. It was learned that the patient enjoyed these symptoms.For this purpose, the patient made a habit of eating a completely salt-free diet and consumed plenty of water. Although pagophagia is considered to cause hyponatremia because it causes excessive water intake, there are not enough cases reported. It is to be noted that people develop habits or addiction to things they like

Development and validation of UV/VIS spectroscopy method for determination of atezolizumab in pharmaceutical products

Aim: The aim of this study was to develop and validate a simple, fast, and reliable UV visible methodology for the determination of atezolizumab in pharmaceutical products. Methods: The maximum wavelength of atezolizumab was determined using a UV/Vis spectrum and the calibration curve has been established. Validation studies were carried out to determine the reliability of the spectrophotometer method used in quantification of pharmaceutical products. Results: According to the experimental data, the developed method was linear in a range varying from 0.10 to 1.50 mg.mL-1 determined by 6 individuals calibrations points. The r2 value was 0.9995 indicating a 99.95% correlation in linearity and precision. The robustness showed good and similar values and the limit of detection and limit of quantification were 0.005 mg.mL-1 and 0.018 mg.mL-1, respectively. Conclusion: The data corroborates the reliability as applicability of the developed UV/Vis spectroscopy method for quantitatively determining the amount of atezolizumab in pharmaceutical products

Preparation and characterization of non-viral gene delivery systems with pEGFP-C1 Plasmid DNA

In recent years, non-viral delivery systems for plasmid DNA have become particularly important. They can overcome the disadvantages of viral systems such as insertional mutagenesis and unpredicted immunogenicity. Some additional advantages of non-viral gene delivery systems are; good stability, low cost, targetability, delivery of a high amount of genetic materials. The aim of the study was to develop novel non-viral nanosystems suitable for gene delivery. Two formulations were developed for this purpose: water-in-oil microemulsion (ME) and solid lipid nanoparticles (SLN). The microemulsion was composed of Peceol, Tween 80, Plurol oleique, ethanol and water. The SLN was consisting of Precirol, Esterquat-1 (EQ1), Tween 80, Lecithin, ethanol and water. Characterization studies were carried out by measuring particle size, zeta potential, viscosity and pH. TEM imaging was performed on SLN formulations. Protection against DNase I degradation was examined. Cytotoxicity and transfection efficacy of selected formulations were tested on L929 mouse fibroblast cells. Particle sizes of complexes were below 100 nm and with high positive zeta potential. TEM images revealed that SLNs are spherical. The SLN:DNA complexes have low toxicity and good transfection ability. All results showed that the developed SLN formulations can be considered as suitable non-viral gene delivery systems

Effects of hospital facilities on patient outcomes after cancer surgery: an international, prospective, observational study

Background Early death after cancer surgery is higher in low-income and middle-income countries (LMICs) compared with in high-income countries, yet the impact of facility characteristics on early postoperative outcomes is unknown. The aim of this study was to examine the association between hospital infrastructure, resource availability, and processes on early outcomes after cancer surgery worldwide.Methods A multimethods analysis was performed as part of the GlobalSurg 3 study-a multicentre, international, prospective cohort study of patients who had surgery for breast, colorectal, or gastric cancer. The primary outcomes were 30-day mortality and 30-day major complication rates. Potentially beneficial hospital facilities were identified by variable selection to select those associated with 30-day mortality. Adjusted outcomes were determined using generalised estimating equations to account for patient characteristics and country-income group, with population stratification by hospital.Findings Between April 1, 2018, and April 23, 2019, facility-level data were collected for 9685 patients across 238 hospitals in 66 countries (91 hospitals in 20 high-income countries; 57 hospitals in 19 upper-middle-income countries; and 90 hospitals in 27 low-income to lower-middle-income countries). The availability of five hospital facilities was inversely associated with mortality: ultrasound, CT scanner, critical care unit, opioid analgesia, and oncologist. After adjustment for case-mix and country income group, hospitals with three or fewer of these facilities (62 hospitals, 1294 patients) had higher mortality compared with those with four or five (adjusted odds ratio [OR] 3.85 [95% CI 2.58-5.75]; p<0.0001), with excess mortality predominantly explained by a limited capacity to rescue following the development of major complications (63.0% vs 82.7%; OR 0.35 [0.23-0.53]; p<0.0001). Across LMICs, improvements in hospital facilities would prevent one to three deaths for every 100 patients undergoing surgery for cancer.Interpretation Hospitals with higher levels of infrastructure and resources have better outcomes after cancer surgery, independent of country income. Without urgent strengthening of hospital infrastructure and resources, the reductions in cancer-associated mortality associated with improved access will not be realised

Investigation of the potential therapeutic effect of cationic lipoplex mediated fibroblast growth factor-2 encoding plasmid DNA delivery on wound healing

Background Developing an alternative and efficient therapy for wound healing has been an important research topic for pharmaceutical sciences. A straightforward but effective system for delivering fibroblast growth factor-2 (FGF-2) encoding plasmid DNA (pFGF-2) for wound healing therapy was aimed to develop in this study. Methods In order to provide the delivery of pFGF-2, a delivery vector, namely, cationic lipid nanopArticle (cLN) was developed by the melt-emulsification process, complexed with pFGF-2 to form a lipoplex system and further characterized. The pFGF-2 binding and protecting ability of lipoplexes were evaluated. The cytotoxicity and transfection efficiency of the lipoplexes, FGF-2 expression levels, and in vitro wound healing ability have been investigated on the L929 fibroblast cell line. Results The obtained lipoplex system has a pArticle size of 88.53 nm with a low PDI (0.185), and zeta potential values of 27.8 mV with a spherical shape. The ability of cLNs to bind pFGF-2 and protect against nucleases was demonstrated by gel retardation assay. Furthermore, the developed FGF-2 carrying lipoplexes system showed significant transfection and FGF-2 expression ability comparing naked plasmid. Finally, scratch assay revealed that the developed system is able to promote in vitro cell proliferation/migration in 48 h. Conclusion Promising results have been achieved with the use of lipoplexes carrying pFGF-2, and this approach could be considered as a potentially applicable concept for the future gene-based wound healing therapies.This project was supported by Izmir Katip Celebi University, Scientific Research Projects Coordinatorship (Project Number: 2019-ONAP-ECZF-0002).Izmir Katip Celebi University, Scientific Research Projects Coordinatorship [2019-ONAP-ECZF-0002

A comparative study of cationic liposomes for gene delivery

In the field of gene delivery, non-viral vectors have become more attractive carriers for nucleic acids since they can overcome the significant drawbacks of viral systems such as safety, immunogenicity, and oncogenicity. Among non-viral vectors, cationic liposome-mediated gene delivery gives promising results for gene therapy approaches. This study aimed to develop cationic liposomes and examine the effectiveness in terms of gene delivery. For this purpose, cholesterol, lecithin, and cationic lipid containing liposomes have been developed by film hydration method. Two different cationic lipids, DDAB and EQ, and their different mole ratios were investigated. Characterization studies showed that obtained liposomes have appropriate physiochemical characteristics (similar to 100 nm, homogenous in size and positive zeta potential) for gene delivery. Gel retardation assay revealed that they have DNA binding and protection ability against nucleases. According to the cytotoxicity evaluation performed on L929 cell line, EQ containing liposomes shows significantly less toxicity comparing DDAB containing liposomes (p<0.05). Furthermore, in vitro transfection study revealed that increasing the EQ mole ratio has increased transfection ability. The stability results showed that the optimal liposome which contains EQ in a higher mole ratio is stable during 90 days at 4 degrees C. Based on these findings, we propose that the developed optimal liposome system in this study could be considered as a suitable nucleic acid delivery vehicle base for gene therapy.TUBITAK (The Scientific and Technological Research Council of Turkey) [218S840]; Izmir Katip Celebi University Research Fund [2020-ODL-ECZF-0001, 108S083]This study has been financially supported by a grant from TUBITAK (The Scientific and Technological Research Council of Turkey) (project no: 218S840) and Izmir Katip Celebi University Research Fund (project no: 2020-ODL-ECZF-0001). We appreciate the project 108S083 (TUBITAK) in which the Malvern Zetasizer Instrument was purchased

Yeni bir manyetik nanopartikül üretim yöntemi ve manyetik alan düzenlemeleri altında manyetofeksiyon

Yeni Bir Manyetik Nanopartikül Üretim Yöntemi ve Manyetik Alan Düzenlemeleri Altında Manyetofeksiyon Amaç: Yeni bir MNP üretim yönteminin geliştirilmesi, elde edilen MNP’lerin karakterizasyonu ve transfeksiyon etkinliğinin manyetik alan düzenlemeleri altında belirlenmesi amaçlandı. Gereç ve Yöntem: Çoklu emülsiyonların mikroreaktörler olarak kullanımı ile MNP üretimi ilk defa tez kapsamında gerçekleştirildi. MNP:DNA kompleksleri oluşturuldu. MNP’lerin partikül boyutu, partikül boyut dağılımı, zeta potansiyeli, manyetizma değerleri ölçüldü, SDS teşvikli DNA serbestleşmesi ve DNaz I enzimine karşı koruma çalışmaları ile organizmada taşıyıcı sistem olarak potansiyelleri değerlendirildi. İstenen özellikleri sağlayan MNP’lerin yüzey özellikleri, manyetik çekirdeği, partikül boyutları DSC, Raman spektroskopisi, SEM ve TEM çalışmalarıyla incelendi, stabilite değerleri araştırıldı. MNP’lere uygulanacak manyetik alanın regülasyonunu sağlamak amacıyla yeni bir cihaz tasarlanarak kullanıldı. Hücre kültüründe belirlenen MNP:DNA komplekslerinin sitotoksisite ve transfeksiyon etkinliği değerlendirildi Bulgular ve Sonuçlar: Metin içerisinde ayrıntılı olarak sunulan sonuçlara göre; transfeksiyon için optimal formülasyon olarak GMS-MNP-1 belirlendi ve karakterize edildi. GMS-MNP-1’in transfeksiyon etkinliği %30,1 olarak bulundu. Pozitif kontrol olarak kullanılan PolyMag® kadar iyi derecede etkili olduğu ve sitotoksik olmadığı gösterildi. Anahtar sözcükler: Manyetofeksiyon, Gen aktarımı, Katyonik nanopartiküller, Çoklu emülsiyon SUMMARY A Novel Magnetic Nanoparticle Production Method and Magnetofection Under Magnetic Field Regulations Aim: It was aimed to develop a novel magnetic nanoparticle production method, characterization of obtained MNPs, and determining their transfection efficiency under magnetic field regulations Materials and Methods: Novel magnetic nanoparticle production method was developed by using multiple emulsions as microreactors for the first time. MNP:DNA complexes were formed. Particle size, polydispersity index, zeta potential and magnetic values of MNPs was measured and their potential of being a delivery system in the organism was evaluated by SDS release, protection against DNase I enzyme studies. MNPs which have desired properties were analyzed for their surface properties, magnetic core material, particle size by DSC, Raman Spectroscopy, SEM and TEM; their stability was also investigated. In order to use regulating the applied magnetic field to the MNPs, a novel device was designed. Cytotoxicity and transfection efficiency of MNP:DNA complexes were evaluated in cell culture. Results and Conclusion: As to the findings that are presented in detail in the text; MNP-GMS-1 were determined as optimal formulation for transfection and were characterized. Transfection efficiency of MNP-GMS-1 was found 30,1%. It was evaluated that MNP-GMS-1 was able to transfect cells as efficiently as positive control-PolyMag® and was not cytotoxic. Keywords: Magnetofection, gene delivery, cationic nanoparticles, multiple emulsio