Perls reaction - histochemistry of iron in liver

Uloga željeza u organizmu je jako bitna i raznolika. Željezo je esencijalni element i možemo ga podijeliti na hemsko i nehemsko. Pojam nehemsko željezo podrazumijeva heterogene vrste željezovih kompleksa u kojima je željezo slabo vezano na organske baze i proteine niske molekulske mase, za razliku od hemskog željeza gdje je taj element vezan na protoporfirinski prsten velike molekulske mase. Za histokemiju nehemskog željeza otopljenog u kiseloj otopini koriste se dvije histokemijske metode: Perlova i Turnbull metoda te i druge metode koje koriste kelatore željeza i ovise o stvaranju netopivih čestica željeza. Najčešće korištena u laboratorijima je Perlova metoda i njome se mogu dokazati i fero i feri ioni željeza, dok je Turnbull metoda specifična samo za fero ione. Međutim, perfuzijom Turnbull metode in vivo može se jasno pokazati distribucija feri željeza, osobito u lizosomima. Za elektronsku mikroskopiju Perlova i Turnbull reakcija se mogu pojačati DAB/ srebro/ zlato metodom da bi vizualizacija željeza bila što bolja, te se primjenjuju i željezo sulfid metoda kao i bojenje redox–aktivnog željeza vodikovim peroksidom ili DAB-om. Unatoč svim prednostima opisanih metoda, kvantitativna mjerenja željeza u pojedinim uzorcima su dosta komplicirana i teško izvediva. Danas se za kvantitativnu procjenu kelatirajućeg Fe2+ najboljom pokazala metoda zasnovana na suzbijanju flourescencije indikatora metala ( PG , SK, rodaina, kalceina, ...) te na njenom ponovnom uspostavljanju poznatim količinama različitih kelatora dvovalentnih metala, opisana u poglavlju 5.2. Sve spomenute metode imaju veliku ulogu u dijagnosticiranju različitih poremećaja i oštećenja tkiva.The role of iron in the body is very important and diverse. Iron is an essential element and we can devide it on hem and nonheme iron. The term nonheme iron includes heterogeneous types of complexes in which iron is loosely bound to low-molecular weight organic bases and proteins, unlike hem iron where this element is linked to the high molecular weight protoporphyrin ring. For histochemistry of nonheme iron dissolved in acid solution , in use are two histochemical methods: Perls and Turnbull method and other methods using iron chelators and dependent on the creation of insoluble particles of iron. The most commonly used in laboratories is a Perls method and it stains ferrous and ferric iron ions, while the Turnbull method is specific only to ferrous ions. However, perfusion Turnbull methods in vivo can clearly demonstrate the distribution of ferrous iron, particularly in the lysosomes. For electron microscopy Perls and Turnbull reactions are enhanced by DAB / silver / gold method for better visualization and iron sulfide staining methods as well as redox-active iron with hydrogen peroxide or DAB-om are also applicable for electon microscopy. Despite all the advantages of the described methods, quantitative measurements of iron in some samples are quite complicated and difficult feasible. Today, for the quantitative assessment of chelating Fe2 + showed the best method based on quenching fluorescence metal indicators (PG, SG, rodaina, kalceina, ...) and its dequenching of the known quantities of various divalent metal chelating treatment, described in section 5.2. All these methods have an important role in the diagnosis of various disorders and tissue damage

Perls reaction - histochemistry of iron in liver

Uloga željeza u organizmu je jako bitna i raznolika. Željezo je esencijalni element i možemo ga podijeliti na hemsko i nehemsko. Pojam nehemsko željezo podrazumijeva heterogene vrste željezovih kompleksa u kojima je željezo slabo vezano na organske baze i proteine niske molekulske mase, za razliku od hemskog željeza gdje je taj element vezan na protoporfirinski prsten velike molekulske mase. Za histokemiju nehemskog željeza otopljenog u kiseloj otopini koriste se dvije histokemijske metode: Perlova i Turnbull metoda te i druge metode koje koriste kelatore željeza i ovise o stvaranju netopivih čestica željeza. Najčešće korištena u laboratorijima je Perlova metoda i njome se mogu dokazati i fero i feri ioni željeza, dok je Turnbull metoda specifična samo za fero ione. Međutim, perfuzijom Turnbull metode in vivo može se jasno pokazati distribucija feri željeza, osobito u lizosomima. Za elektronsku mikroskopiju Perlova i Turnbull reakcija se mogu pojačati DAB/ srebro/ zlato metodom da bi vizualizacija željeza bila što bolja, te se primjenjuju i željezo sulfid metoda kao i bojenje redox–aktivnog željeza vodikovim peroksidom ili DAB-om. Unatoč svim prednostima opisanih metoda, kvantitativna mjerenja željeza u pojedinim uzorcima su dosta komplicirana i teško izvediva. Danas se za kvantitativnu procjenu kelatirajućeg Fe2+ najboljom pokazala metoda zasnovana na suzbijanju flourescencije indikatora metala ( PG , SK, rodaina, kalceina, ...) te na njenom ponovnom uspostavljanju poznatim količinama različitih kelatora dvovalentnih metala, opisana u poglavlju 5.2. Sve spomenute metode imaju veliku ulogu u dijagnosticiranju različitih poremećaja i oštećenja tkiva.The role of iron in the body is very important and diverse. Iron is an essential element and we can devide it on hem and nonheme iron. The term nonheme iron includes heterogeneous types of complexes in which iron is loosely bound to low-molecular weight organic bases and proteins, unlike hem iron where this element is linked to the high molecular weight protoporphyrin ring. For histochemistry of nonheme iron dissolved in acid solution , in use are two histochemical methods: Perls and Turnbull method and other methods using iron chelators and dependent on the creation of insoluble particles of iron. The most commonly used in laboratories is a Perls method and it stains ferrous and ferric iron ions, while the Turnbull method is specific only to ferrous ions. However, perfusion Turnbull methods in vivo can clearly demonstrate the distribution of ferrous iron, particularly in the lysosomes. For electron microscopy Perls and Turnbull reactions are enhanced by DAB / silver / gold method for better visualization and iron sulfide staining methods as well as redox-active iron with hydrogen peroxide or DAB-om are also applicable for electon microscopy. Despite all the advantages of the described methods, quantitative measurements of iron in some samples are quite complicated and difficult feasible. Today, for the quantitative assessment of chelating Fe2 + showed the best method based on quenching fluorescence metal indicators (PG, SG, rodaina, kalceina, ...) and its dequenching of the known quantities of various divalent metal chelating treatment, described in section 5.2. All these methods have an important role in the diagnosis of various disorders and tissue damage

Perls reaction - histochemistry of iron in liver

Uloga željeza u organizmu je jako bitna i raznolika. Željezo je esencijalni element i možemo ga podijeliti na hemsko i nehemsko. Pojam nehemsko željezo podrazumijeva heterogene vrste željezovih kompleksa u kojima je željezo slabo vezano na organske baze i proteine niske molekulske mase, za razliku od hemskog željeza gdje je taj element vezan na protoporfirinski prsten velike molekulske mase. Za histokemiju nehemskog željeza otopljenog u kiseloj otopini koriste se dvije histokemijske metode: Perlova i Turnbull metoda te i druge metode koje koriste kelatore željeza i ovise o stvaranju netopivih čestica željeza. Najčešće korištena u laboratorijima je Perlova metoda i njome se mogu dokazati i fero i feri ioni željeza, dok je Turnbull metoda specifična samo za fero ione. Međutim, perfuzijom Turnbull metode in vivo može se jasno pokazati distribucija feri željeza, osobito u lizosomima. Za elektronsku mikroskopiju Perlova i Turnbull reakcija se mogu pojačati DAB/ srebro/ zlato metodom da bi vizualizacija željeza bila što bolja, te se primjenjuju i željezo sulfid metoda kao i bojenje redox–aktivnog željeza vodikovim peroksidom ili DAB-om. Unatoč svim prednostima opisanih metoda, kvantitativna mjerenja željeza u pojedinim uzorcima su dosta komplicirana i teško izvediva. Danas se za kvantitativnu procjenu kelatirajućeg Fe2+ najboljom pokazala metoda zasnovana na suzbijanju flourescencije indikatora metala ( PG , SK, rodaina, kalceina, ...) te na njenom ponovnom uspostavljanju poznatim količinama različitih kelatora dvovalentnih metala, opisana u poglavlju 5.2. Sve spomenute metode imaju veliku ulogu u dijagnosticiranju različitih poremećaja i oštećenja tkiva.The role of iron in the body is very important and diverse. Iron is an essential element and we can devide it on hem and nonheme iron. The term nonheme iron includes heterogeneous types of complexes in which iron is loosely bound to low-molecular weight organic bases and proteins, unlike hem iron where this element is linked to the high molecular weight protoporphyrin ring. For histochemistry of nonheme iron dissolved in acid solution , in use are two histochemical methods: Perls and Turnbull method and other methods using iron chelators and dependent on the creation of insoluble particles of iron. The most commonly used in laboratories is a Perls method and it stains ferrous and ferric iron ions, while the Turnbull method is specific only to ferrous ions. However, perfusion Turnbull methods in vivo can clearly demonstrate the distribution of ferrous iron, particularly in the lysosomes. For electron microscopy Perls and Turnbull reactions are enhanced by DAB / silver / gold method for better visualization and iron sulfide staining methods as well as redox-active iron with hydrogen peroxide or DAB-om are also applicable for electon microscopy. Despite all the advantages of the described methods, quantitative measurements of iron in some samples are quite complicated and difficult feasible. Today, for the quantitative assessment of chelating Fe2 + showed the best method based on quenching fluorescence metal indicators (PG, SG, rodaina, kalceina, ...) and its dequenching of the known quantities of various divalent metal chelating treatment, described in section 5.2. All these methods have an important role in the diagnosis of various disorders and tissue damage

Association of the Human Leukocyte Antigen region with renal carcinoma

U ovom radu analizirani su geni i aleli HLA-A, -B i -DRB1 te aleli mikrosatelita TNFa, TNFb i TNFd među 62 nesrodna bolesnika s karcinomom bubrega. Utvrđeno je da nema statistički značajne razlike u učestalosti alela i gena HLA između bolesničke i kontrolne skupine. Podjelom bolesničke skupine s obzirom na veličinu i gradus tumora te zahvaćenost limfnih čvorova tumorom, uočene razlike u učestalosti gena HLA-B*27 (p=0,0073), -DRB1*12 (p=0,0492) i -B*08 (p=0,0441) gube se korigiranjem vrijednosti p, dok bolesnici s karcinomom gradusa 4 pokazuju povećanu učestalost gena HLA-DRB*13 (p=0,0394) i nakon korigiranja u odnosu na kontrolu i skupine bolesnika sa gradusima 2 i 3. Nije uočena razlika u učestalosti alela TNFa i TNFd u skupini bolesnika u usporedbi s kontrolom za razliku od alela TNFb kod kojeg je uočena značajno veća učestalost TNFb-5 (p=3,7x10-7), te smanjena učestalost TNFb-3 (p=0,0002) kod bolesnika. Također TNFb-5 se pojavljuje češće kod bolesnika s tumorom >7 cm i sa gradusom 4, dok je učestalost TNFb-3 u obje skupine značajno smanjena u odnosu na kontrolu. Iz uočenih razlika bi se TNFb-5 mogao smatrati alelom podložnosti, a TNFb-3 zaštitnim alelom u nastanku karcinoma bubrega.In this study, we analyzed HLA-A, -B and -DRB1 genes and alleles and TNFa, TNFb and TNFd microsatellite alleles among 62 unrelated patients with renal carcinoma. No statistically significant difference was found in HLA genes and alleles frequency among patients with renal carcinoma in comparison to controls. Comparison which took into account size, tumor gradus and lymph node metastasis, revealed differences in the frequency of HLA-B*27 (p=0.0073), -DRB1*12 (p=0.0492) and -B*08 (p=0.0441) but only before the p value correction. Patients with gradus 4 showed a significant increase of HLA-DRB1*13 frequency (p=0.0394) in comparison to controls and patients with gradus 2 and 3 even after p value correction. There was no difference in the frequency of TNFa and TNFd alleles among patients in comparison to controls as opposed to TNFb locus for which a higher frequency of TNFb-5 (p=3.7X10-7), and reduced frequency of TNFb-3 (p=0.0002) among patients was observed. TNFb-5 allele occurs more frequently in patients with tumors >7 cm and gradus 4, while the frequency of TNFb-3 in both subgroups shows a decrease compared to controls. The observed difference leads to conclusion that TNFb-5 could infer susceptibility and TNFb-3 protection in the development of renal carcinoma

Tumor Organoid and Spheroid Models for Cervical Cancer

Cervical cancer is one of the most common malignant diseases in women worldwide. Despite the global introduction of a preventive vaccine against the leading cause of cervical cancer, human papillomavirus (HPV) infection, the incidence of this malignant disease is still very high, especially in economically challenged areas. New advances in cancer therapy, especially the rapid development and application of different immunotherapy strategies, have shown promising pre-clinical and clinical results. However, mortality from advanced stages of cervical cancer remains a significant concern. Precise and thorough evaluation of potential novel anti-cancer therapies in pre-clinical phases is indispensable for efficient development of new, more successful treatment options for cancer patients. Recently, 3D tumor models have become the gold standard in pre-clinical cancer research due to their capacity to better mimic the architecture and microenvironment of tumor tissue as compared to standard two-dimensional (2D) cell cultures. This review will focus on the application of spheroids and patient-derived organoids (PDOs) as tumor models to develop novel therapies against cervical cancer, with an emphasis on the immunotherapies that specifically target cancer cells and modulate the tumor microenvironment (TME)

Towards Novel Gene and Cell Therapy Approaches for Cervical Cancer

Cervical cancer is one of the most common malignancies in women, and the majority of cases are caused by infection with high-risk human papilloma virus (HPV) subtypes. Despite effective preventative measures, such as vaccinations against HPV, over 300,000 women die world-wide from cervical cancer each year. Once cervical cancer is diagnosed, treatment may consist of radial hysterectomy, or chemotherapy and radiotherapy, or a combination of therapies dependent upon the disease stage. Unfortunately, overall prognosis for patients with metastatic or recurrent disease remains poor. In these cases, immunotherapies may be useful based on promising preclinical work, some of which has been successfully translated to the clinic. For example, approaches using monoclonal antibodies directed against surface proteins important for control of immune checkpoints (i.e., immune checkpoint inhibitors) were shown to improve outcome in many cancer settings, including cervical cancer. Additionally, initial clinical studies showed that application of cytotoxic immune cells modified to express chimeric antigen receptors (CAR) or T cell receptors (TCR) for better recognition and elimination of tumor cells may be useful to control cervical cancer. This review explores these important topics, including strengths and limitations of standard and developing approaches, and how some novel treatment strategies may be optimally used to offer the best possible treatment for cervical cancer patients

The M25 gene products are critical for the cytopathic effect of mouse cytomegalovirus

Abstract Cell rounding is a hallmark of the cytopathic effect induced by cytomegaloviruses. By screening a panel of deletion mutants of mouse cytomegalovirus (MCMV) a mutant was identified that did not elicit cell rounding and lacked the ability to form typical plaques. Altered cell morphology was assigned to the viral M25 gene. We detected an early 2.8 kb M25 mRNA directing the synthesis of a 105 kDa M25 protein, and confirmed that a late 3.1 kb mRNA encodes a 130 kDa M25 tegument protein. Virions lacking the M25 tegument protein were of smaller size because the tegument layer between capsid and viral envelope was reduced. The ΔM25 mutant did not provoke the rearrangement of the actin cytoskeleton observed after wild-type MCMV infection, and isolated expression of the M25 proteins led to cell size reduction, confirming that they contribute to the morphological changes. Yields of progeny virus and cell-to-cell spread of the ΔM25 mutant in vitro were diminished and replication in vivo was impaired. The identification of an MCMV gene involved in cell rounding provides the basis for investigating the role of this cytopathic effect in CMV pathogenesis

CAR-NK Cells Targeting HER1 (EGFR) Show Efficient Anti-Tumor Activity against Head and Neck Squamous Cell Carcinoma (HNSCC)

(1) Background: HNSCC is a highly heterogeneous and relapse-prone form of cancer. We aimed to expand the immunological tool kit against HNSCC by conducting a functional screen to generate chimeric antigen receptor (CAR)-NK-92 cells that target HER1/epidermal growth factor receptor (EGFR). (2) Methods: Selected CAR-NK-92 cell candidates were tested for enhanced reduction of target cells, CD107a expression and IFNγ secretion in different co-culture models. For representative HNSCC models, patient-derived primary HNSCC (pHNSCC) cell lines were generated by employing an EpCAM-sorting approach to eliminate the high percentage of non-malignant cells found. (3) Results: 2D and 3D spheroid co-culture experiments showed that anti-HER1 CAR-NK-92 cells effectively eliminated SCC cell lines and primary HNSCC (pHNSCC) cells. Co-culture of tumor models with anti-HER1 CAR-NK-92 cells led to enhanced degranulation and IFNγ secretion of NK-92 cells and apoptosis of target cells. Furthermore, remaining pHNSCC cells showed upregulated expression of putative cancer stem cell marker CD44v6. (4) Conclusions: These results highlight the promising potential of CAR-NK cell therapy in HNSCC and the likely necessity to target multiple tumor-associated antigens to reduce currently high relapse rates