Spectrophotometric procedures for the rapid characterization of propolis have been performed on p... more Spectrophotometric procedures for the rapid characterization of propolis have been performed on propolis samples from different regions of Croatia. In order to determine the major groups of bioactive compounds in propolis, the following optimised and validated spectrophotometric methods were carried out: the Folin-Ciocalteu method for the content of total phenolics (TPs) and two distinct methods for the content of total flavonoids: aluminium chloride (AlCl 3) complexation method for total flavones/flavonols (TFFs) and the 2.4-dinitrophenylhydrazine (2.4-DNPH) method for total flavanones/dihydroflavonols (TFDs). Validation parameters, including linearity, sensitivity, range, accuracy, limit of detection, limit of quantification, precision and robustness were implemented. The following polyphenol standards were used for the validation procedure: gallic acid (GA), pinocembrin (PC), galangin (GN), quercetin (QC) and a mixture of PC and GN. Validated methods were applied to analyse six samples of raw propolis from Croatian continental and Adriatic regions. The high qualitative/quantitative variability of the TP, TFF and TFD content was observed. Although the method of extraction (ultrasonic-assisted extraction or microwave assisted extraction) showed a non-significant effect on extraction yield (P>0.05) and the polyphenolic concentrations obtained of each sample in general, ultrasonic extraction was found to be more selective. Furthermore, the calibration compound used for constructing the calibration curve highly influenced the final concentrations of TPs and TFFs. The study showed good linearity, accuracy, repeatability, intermediate precision, LOD and LOQ for all three spectrophotometric methods, considering that these analyses are the basis for further research into the individual polyphenolic compounds in the propolis samples covered by this research.
Standard cancer therapies commonly require multiple agents. However, use of multiple agents contr... more Standard cancer therapies commonly require multiple agents. However, use of multiple agents contributes to increased toxicity resulting in poor treatment outcome. Thus, combined chemotherapy must be optimized to increase tumor response and at the same time lower its toxicity. Antioxidants have been shown to enhance the effect of certain chemotherapeutic agents on tumor cells in vitro and in vivo. However, this effect differs depending upon the type of tumor and the drugs. The objective of this study was to see whether pretreatment with antioxidant mixture could enhance the cytotoxic and apoptotic effect of commonly used chemotherapeutic agents, irinotecan, cisplatin, doxorubicin and epirubicin for the treatment of Echrlich ascites tumor or of mammary carcinoma in mice. Chemotherapeutic agents are known to induce nuclear factor KB (NF- B) activity in tumor cells, resulting in lower cell killing and drug resistance. Collectively, our studies and those by others showed that different polyphenolic/flavonoid components separated from propolis as well as propolis alone could „ sensitize“ cancer cells to chemotherapeutic agents, likely by blocking NF- B activation induced by chemotherapeutic agents both in vitro and in vivo. These results provide experimental evidence showing that novel therapeutic strategies for human cancer could be developed to achieve better treatment outcome by introducing flavonoids in the therapeutic regimen. Positive outcome of combined treatment with propolis and chemotherapeutics was expressed by the influence of these treatments on reduced cytotoxicity to peripheral blood leukocytes, liver and kidney cells.Therefore, polyphenolic/flavonoid components could be potentially useful in combination with chemotherapeutic agents for the treatment of different types of human cancer.
Thermal imaging has been utilised, both preclinically and clinically, as a tool for assessing inf... more Thermal imaging has been utilised, both preclinically and clinically, as a tool for assessing inflammation. Psoriasis is a chronic inflammatory skin disease characterised by hyperkeratosis, dermal inflammatory infiltrate and increased angiogenesis. The aim of the present study was to assess the usefulness of thermography in psoriatic lesion regression after topical treatment with bee propolis, recognised as potent antioxidants and anti-inflammatory agents. We monitored the inflammation process induced by irritants such as n-Hexyl salycilate (HXS) or Di-n-Propyl Disulfide (PPD) by histopatological assessment of the skin, thermographic scanning, total number of inflammatory cells in the peritoneal cavity, differential analysis of cells in the peritoneal cavity, macrophage spreading index, haematological and biochemical parameters, frequencies of micronucleated reticulocytes, lipid peroxidation and glutathione assay in the skin. Topically applied ethanolic extract of propolis (EEP) with HXS or PPD reduced the lipid peroxidation in the skin and total number of inflammatory cells in the skin and peritoneal cavity, functional activity of macrophages, the number of micronuclei in mouse peripheral blood reticulocytes and enzymatic activity of ALP and AST. These results demonstrate that topical application of EEP may improve psoriatic-like skin lesions by suppressing functional activity of macrophages and ROS production. Taken together, it is suggested that EEP can safely be utilised in the prevention of psoriasis-related inflammatory changes without causing any toxic effect.
Oxidative stress may lead to extensive bone loss and skeletal fragility, two characteristics of o... more Oxidative stress may lead to extensive bone loss and skeletal fragility, two characteristics of osteoporosis. We studied the bone‐protective effect of quercetin (QU) in the retinoic acid induced model of osteoporosis (RMO) and in healthy rats. Densitometry, osteocalcin (OC), Cterminal cross‐linked telopeptide (CTX), calcium (Ca) and phosphorus (P) content of bone femur, geometric measurement on the femur, histology, hematological and biochemical parameters as well as lipid peroxidation (MDA) and glutathione levels (GSH) of liver and kidney cells were applied to all groups. Retinoic acid administration (80 mg kg‐1) revealed significant increase in biochemical parameters, MDA level, OC and CTX values and a significant decrease in values of BMD, Ca and P in the femur and GSH levels in liver and kidney compared to control. Administration of QU intragastrically (100 mg kg‐1 bw) daily concomitant with retinoic acid resulted in significantly reduction of CTX values and MDA in comparison to RMO. QU also exerted positive effects on biochemical parameters, values of Ca and P, BMD values in distal part, femoral weight and length and the GSH level. Concomitant treatments of rats with retinoic acid and alendronate had toxic effect to animals and resulted in a significant decrease in body mass of 5.67% after two weeks. The protective role of the QU against the toxicity of retinoic acid‐induced osteoporosis in rats gives a hope that QU may have similar protective action in humans.
The aim of this study was to detect changes in the concentration of serum immunoglobulins followi... more The aim of this study was to detect changes in the concentration of serum immunoglobulins following vaccination against classical swine fever (CSF) with an attenuated C strain and a subunit E2 vaccine. Furthermore, the adjuvanticity of an attenuated parapoxvirus ORF virus for the subunit vaccine against CSF was evaluated. Peripheral blood samples were collected before the vaccination and at post-vaccination days 4, 10, 21 and 28. The samples were assessed by a colorimetric method for the detection of total proteins, as well as albumin, IgA and IgM levels and by radial immunodiffusion to record the IgG level. Our findings are in accordance with the normal concentrations of porcine IgG, IgA and IgM. However, a significant increase of some immunoglobulin classes was recorded. The increase of the IgM level in vaccinated pigs confirmed an early development of humoral immunity. Interestingly, the subunit E2 vaccine induced the increase of IgM earlier then did the attenuated C strain. Since the IgG concentratiom was not significantly increased we assumed that the period of 28 days following vaccination was too short to detect any changes in the IgG level, thus reflecting a late humoral immune response. Although, IgA antibodies are mostly responsible for humoral immunity at the mucosal surfaces, in our experiment the attenuated C strain induced a significantly higher production of this immunoglobulin class in the serum very early (on day 4) following vaccination. This could be ascribed to the affinity of IgA antibodies to neutralize or agglutinate virus particles. Early appearance (4 and 10 days after the vaccination) of a significantly higher concentration of IgG and IgM could be induced by the ORF virus strain D1701 applied as an adjuvant.
Thermal imaging has been utilised, both preclinically and clinically, as a tool for assessing inf... more Thermal imaging has been utilised, both preclinically and clinically, as a tool for assessing inflammation. Psoriasis is a chronic inflammatory skin disease characterised by hyperkeratosis, dermal inflammatory infiltrate and increased angiogenesis. The aim of the present study was to assess the usefulness of thermography in psoriatic lesion regression after topical treatment with bee propolis, recognised as potent antioxidants and anti-inflammatory agents. We monitored the inflammation process induced by irritants such as n-Hexyl salycilate (HXS) or Di-n-Propyl Disulfide (PPD) by histopatological assessment of the skin, thermographic scanning, total number of inflammatory cells in the peritoneal cavity, differential analysis of cells in the peritoneal cavity, macrophage spreading index, haematological and biochemical parameters, frequencies of micronucleated reticulocytes, lipid peroxidation and glutathione assay in the skin. Topically applied ethanolic extract of propolis (EEP) with HXS or PPD reduced the lipid peroxidation in the skin and total number of inflammatory cells in the skin and peritoneal cavity, functional activity of macrophages, the number of micronuclei in mouse peripheral blood reticulocytes and enzymatic activity of ALP and AST. These results demonstrate that topical application of EEP may improve psoriatic-like skin lesions by suppressing functional activity of macrophages and ROS production. Taken together, it is suggested that EEP can safely be utilised in the prevention of psoriasis-related inflammatory changes without causing any toxic effect.
In this paper we report for the first time a method for the extraction of the protein fraction fr... more In this paper we report for the first time a method for the extraction of the protein fraction from Feijoa sellowiana Berg. fruit and its electrophoretic analysis. In addition we analyzed the protein fraction for its antioxidant activity and its effectiveness against different Gram-positive and Gram-negative bacteria both as American type culture collection (ATCC) standard and clinically isolated strains. Both antimicrobial and antioxidant activity resulted stronger respect to that previously obtained for the acetonic extract from the same fruit. Further, we study both activities also on the product of in vitro gastrointestinal digestion of F. sellowiana fruit proteins. The results showed that in vitro gastrointestinal digestion increased 10 fold the antioxidant activity, while the antimicrobial activity, tested only on ATCC strains, resulted from 2 to 4 fold increased. MTT assays showed the nontoxicity of these proteins both before and after digestion.
This study was carried out to find whether propolis and its polyphenolic/flavonoids compounds may... more This study was carried out to find whether propolis and its polyphenolic/flavonoids compounds may induce cytotoxicity in primary culture of human urinary bladder transitional cell carcinoma (TCC) cells as compared to normal urinary bladder epithelial cells. Pieces of TCC or normal epithelial tissue were collected by transurethral surgery from patients in different stages (grade G1,G2, G3) of TCC. Incubation of TCC cells for cytotoxicity testing were carried out with or without diffrent concentration (50, 150, 300 µg/ml) of test components. The cytotoxicity of two preparation of propolis (water and ethanolic extract of propolis; WSDP or EEP) and its polyphenolic compounds (caffeic acid, naringin, chrysin, quercetin) was determined using trypan blue exlusion assay. Findings suggest that EEP is the most effective in inhibition of urinary bladder TCC cell proliferation as compared to WSDP or single flavonoids derived from propolis. All test components showed no cytotoxicity to normal epithelial cells. The result of this study may provide great impact on the potential activity of EEP as an adjuvant to surgery, to suppress or prevent tumor recurrence in urinary bladder since only a few anti-cancer drugs have been effective in tumor control. Since immunomodulation by BCG has been used to impruve the results of surgery it is likely that propolis preparation (EEP) as immunomodulating compound may be a substitute for mycobacterial treatment since propolis preparation or its polyphenolic components have expressed no side effect after treatment in animal models.
The aim of this study was to evaluate the in vitro influence of mitogens on the lymphoproliferati... more The aim of this study was to evaluate the in vitro influence of mitogens on the lymphoproliferative response after vaccination against classical swine fever (CSF). We also compared the effect of the ORF virus strain D1701, whose immunostimulatory effects in vivo are known, as well as the CSF virus C strain on lymphocyte cell cultures of vaccinated and control pigs. Peripheral blood leucocytes (PBL) were used in a lymphocyte proliferation assay by Concanavalin A, phytohemagglutinin, pokeweed mitogen, the attenuated ORF virus strain D1701 and the CSF virus China (C) strain. PBL were obtained from pigs vaccinated with different CSF vaccines (a subunit gp E2 vaccine, a combination of a subunit vaccine and an ORF virus strain D1701 and attenuated C strain vaccine) and from non-vaccinated pigs. Blood samples for the mitogenic response assay were collected before (day 0), and on the 4th, 10th, 21st and 28th post-vaccination day. PBL of pigs that received the C strain and ORF virus showed in vitro a weaker proliferative response to the mitogens than PBL of pigs vaccinated with the subunit CSF vaccine. The lymphocyte proliferation was observed only in pigs vaccinated with the C strain vaccine after exposure to the same strain in vitro. However, the ORF virus, in the applied concentration did not induce a significant reaction of lymphocytes that originated from either vaccinated or control animals. The results of our study suggest that the applied mitogens may induce in vitro reactivity which could play a significant immunological role after CSF vaccination. In contrary, the attenuated virus strains used as stimulators, induced no significant lymphoproliferation.
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
Using a Swiss albino mouse model of Ehrlich ascites tumor (EAT), we investigated antitumor effica... more Using a Swiss albino mouse model of Ehrlich ascites tumor (EAT), we investigated antitumor efficacy of two preparations of propolis (water and ethanolic extract) or quercetin, a compound of propolis, combined with chemotherapeutic drugs (Cisplatin and Doxorubicin). Egzamination was directed to suppression of tumor progression expressed as the percent increased life span. Propolis or quercetin were administrated ip to animals at doses of 50 mg kg-1 body weight for three consecutive days before tumor inoculation. Doxorubicin (20 mg kg-1) or cisplatin (10 mg kg-1) were given to mice two days after tumor cell inoculation. The results of log rank test between treated and control mice showed:WSDP, p=0.00291 ; WSDP + CIS, p=0.00781 ; WSDP + DOX, p=0.4998 ; EEP, p=0.00902 ; EEP + CIS, p=0.27603 ; EEP + DOX, p=0.22014 ; QU, p=0.27454 ; QU + CIS, p=0.72713 ; QU + DOX, p= 0.02667 ; CIS, p=0.00350 ; DOX, p= 0.2593. Combined treatment with test compounds and Cisplatin resulted in pozitive outcome (p=0, 00853 ; p=0, 00767 ; p=0, 02500) versus cisplatin alone, however combined treatment with doxorubicin was ineffective. Our findings indicate that WSDP, EEP, and quercetin significantly enhanced the antiproliferative and cytotoxic action of cisplatin and that propolis compounds may be usefully used for lowering the toxicity and other deteriorating activities chemotherapeutic drugs.
Kolibaciloza je glavni uzrok oboljevanja i mortaliteta odbijene prasadi. Najcesce je posljedica e... more Kolibaciloza je glavni uzrok oboljevanja i mortaliteta odbijene prasadi. Najcesce je posljedica enterotoksigenih sojeva bakterije E. coli (ETEC) koji proizvode enterotoksine sto ostecuju stijenku crijeva, poticu lucenje sluzi i elektrolita i tako uzrokuju dijareju. Vakcinacija protiv kolibaciloze odbijene prasadi nije u siroj uporabi kao profilakticka mjera. Naime, pokusaji da se potakne specificni imunosni odgovor protiv ETEC sojeva na mukoznim povrsinama crijeva nakon oralne imunizacije s fimbrijskim vakcinama bili su neucinkoviti. Zastita od mikrobnih antigena, kao sto su ETEC sojevi, na mukoznim povrsinama mogla bi se postici indukcijom i regulacijom specificnih obrambenih mehanizama posredovanim imunosnim stanicama u limfatickom tkivu probavnog sustava (LTPS). Vrednovali smo imunogenost F4ac^+ ne-ETEC soja kao izvora replicirajuceg antigena u specificnoj imunizaciji odbijene prasadi temeljem kvalitativnih i kvantitativnih pokazatelja aktivacije njihovih imunosnih stanica u mezenterijskom limfnom cvoru (MLC) uporabom protocne citometrije i imunohistologije, kao i in vitro određivanjem fagocitnih funkcija peritonejskih makrofaga. Sedam dana nakon intragastricke imunizacije sa 10^10 CFU/ml F4ac^+ ne-ETEC soja, analizirali smo CD^+/SWC^+ imunofenotipove stanica iz MLC uporabom monoklonskih protutijela specificnih za CD3, CD8, CD4, CD21, CD14, SWC3 i MHC II povrsinske stanicne molekule. U MLC imunizirane prasadi utvrdili smo porast udjela CD4^+CD8^+ memorijskih stanica, aktiviranih CD25 T pomocnickih i T citotoksicnih limfocita, B limfocita i stanica koje prikazuju antigen. Peritonejski makrofagi imunizirane prasadi imali su vecu sposobnost ingestije cestica. Izrazitija razlika u brojnosti aktiviranih CD25^+ T-limfocita vidljiva je u MLC imunizirane u odnosu na kontrolnu prasad. Prema ovim rezultatima, pokusni vakcinalni F4ac^+ ne-ETEC soj uporabljen za poticanje specificne imunosti odbijene prasadi protiv kolibaciloze izazvao je odgovore mijeloidnih i limfoidnih stanica u odjeljku LTPS tj. u MLC, i unutar peritonejske supljine odbijene prasadi.
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
Spectrophotometric procedures for the rapid characterization of propolis have been performed on p... more Spectrophotometric procedures for the rapid characterization of propolis have been performed on propolis samples from different regions of Croatia. In order to determine the major groups of bioactive compounds in propolis, the following optimised and validated spectrophotometric methods were carried out: the Folin-Ciocalteu method for the content of total phenolics (TPs) and two distinct methods for the content of total flavonoids: aluminium chloride (AlCl 3) complexation method for total flavones/flavonols (TFFs) and the 2.4-dinitrophenylhydrazine (2.4-DNPH) method for total flavanones/dihydroflavonols (TFDs). Validation parameters, including linearity, sensitivity, range, accuracy, limit of detection, limit of quantification, precision and robustness were implemented. The following polyphenol standards were used for the validation procedure: gallic acid (GA), pinocembrin (PC), galangin (GN), quercetin (QC) and a mixture of PC and GN. Validated methods were applied to analyse six samples of raw propolis from Croatian continental and Adriatic regions. The high qualitative/quantitative variability of the TP, TFF and TFD content was observed. Although the method of extraction (ultrasonic-assisted extraction or microwave assisted extraction) showed a non-significant effect on extraction yield (P>0.05) and the polyphenolic concentrations obtained of each sample in general, ultrasonic extraction was found to be more selective. Furthermore, the calibration compound used for constructing the calibration curve highly influenced the final concentrations of TPs and TFFs. The study showed good linearity, accuracy, repeatability, intermediate precision, LOD and LOQ for all three spectrophotometric methods, considering that these analyses are the basis for further research into the individual polyphenolic compounds in the propolis samples covered by this research.
Standard cancer therapies commonly require multiple agents. However, use of multiple agents contr... more Standard cancer therapies commonly require multiple agents. However, use of multiple agents contributes to increased toxicity resulting in poor treatment outcome. Thus, combined chemotherapy must be optimized to increase tumor response and at the same time lower its toxicity. Antioxidants have been shown to enhance the effect of certain chemotherapeutic agents on tumor cells in vitro and in vivo. However, this effect differs depending upon the type of tumor and the drugs. The objective of this study was to see whether pretreatment with antioxidant mixture could enhance the cytotoxic and apoptotic effect of commonly used chemotherapeutic agents, irinotecan, cisplatin, doxorubicin and epirubicin for the treatment of Echrlich ascites tumor or of mammary carcinoma in mice. Chemotherapeutic agents are known to induce nuclear factor KB (NF- B) activity in tumor cells, resulting in lower cell killing and drug resistance. Collectively, our studies and those by others showed that different polyphenolic/flavonoid components separated from propolis as well as propolis alone could „ sensitize“ cancer cells to chemotherapeutic agents, likely by blocking NF- B activation induced by chemotherapeutic agents both in vitro and in vivo. These results provide experimental evidence showing that novel therapeutic strategies for human cancer could be developed to achieve better treatment outcome by introducing flavonoids in the therapeutic regimen. Positive outcome of combined treatment with propolis and chemotherapeutics was expressed by the influence of these treatments on reduced cytotoxicity to peripheral blood leukocytes, liver and kidney cells.Therefore, polyphenolic/flavonoid components could be potentially useful in combination with chemotherapeutic agents for the treatment of different types of human cancer.
Thermal imaging has been utilised, both preclinically and clinically, as a tool for assessing inf... more Thermal imaging has been utilised, both preclinically and clinically, as a tool for assessing inflammation. Psoriasis is a chronic inflammatory skin disease characterised by hyperkeratosis, dermal inflammatory infiltrate and increased angiogenesis. The aim of the present study was to assess the usefulness of thermography in psoriatic lesion regression after topical treatment with bee propolis, recognised as potent antioxidants and anti-inflammatory agents. We monitored the inflammation process induced by irritants such as n-Hexyl salycilate (HXS) or Di-n-Propyl Disulfide (PPD) by histopatological assessment of the skin, thermographic scanning, total number of inflammatory cells in the peritoneal cavity, differential analysis of cells in the peritoneal cavity, macrophage spreading index, haematological and biochemical parameters, frequencies of micronucleated reticulocytes, lipid peroxidation and glutathione assay in the skin. Topically applied ethanolic extract of propolis (EEP) with HXS or PPD reduced the lipid peroxidation in the skin and total number of inflammatory cells in the skin and peritoneal cavity, functional activity of macrophages, the number of micronuclei in mouse peripheral blood reticulocytes and enzymatic activity of ALP and AST. These results demonstrate that topical application of EEP may improve psoriatic-like skin lesions by suppressing functional activity of macrophages and ROS production. Taken together, it is suggested that EEP can safely be utilised in the prevention of psoriasis-related inflammatory changes without causing any toxic effect.
Oxidative stress may lead to extensive bone loss and skeletal fragility, two characteristics of o... more Oxidative stress may lead to extensive bone loss and skeletal fragility, two characteristics of osteoporosis. We studied the bone‐protective effect of quercetin (QU) in the retinoic acid induced model of osteoporosis (RMO) and in healthy rats. Densitometry, osteocalcin (OC), Cterminal cross‐linked telopeptide (CTX), calcium (Ca) and phosphorus (P) content of bone femur, geometric measurement on the femur, histology, hematological and biochemical parameters as well as lipid peroxidation (MDA) and glutathione levels (GSH) of liver and kidney cells were applied to all groups. Retinoic acid administration (80 mg kg‐1) revealed significant increase in biochemical parameters, MDA level, OC and CTX values and a significant decrease in values of BMD, Ca and P in the femur and GSH levels in liver and kidney compared to control. Administration of QU intragastrically (100 mg kg‐1 bw) daily concomitant with retinoic acid resulted in significantly reduction of CTX values and MDA in comparison to RMO. QU also exerted positive effects on biochemical parameters, values of Ca and P, BMD values in distal part, femoral weight and length and the GSH level. Concomitant treatments of rats with retinoic acid and alendronate had toxic effect to animals and resulted in a significant decrease in body mass of 5.67% after two weeks. The protective role of the QU against the toxicity of retinoic acid‐induced osteoporosis in rats gives a hope that QU may have similar protective action in humans.
The aim of this study was to detect changes in the concentration of serum immunoglobulins followi... more The aim of this study was to detect changes in the concentration of serum immunoglobulins following vaccination against classical swine fever (CSF) with an attenuated C strain and a subunit E2 vaccine. Furthermore, the adjuvanticity of an attenuated parapoxvirus ORF virus for the subunit vaccine against CSF was evaluated. Peripheral blood samples were collected before the vaccination and at post-vaccination days 4, 10, 21 and 28. The samples were assessed by a colorimetric method for the detection of total proteins, as well as albumin, IgA and IgM levels and by radial immunodiffusion to record the IgG level. Our findings are in accordance with the normal concentrations of porcine IgG, IgA and IgM. However, a significant increase of some immunoglobulin classes was recorded. The increase of the IgM level in vaccinated pigs confirmed an early development of humoral immunity. Interestingly, the subunit E2 vaccine induced the increase of IgM earlier then did the attenuated C strain. Since the IgG concentratiom was not significantly increased we assumed that the period of 28 days following vaccination was too short to detect any changes in the IgG level, thus reflecting a late humoral immune response. Although, IgA antibodies are mostly responsible for humoral immunity at the mucosal surfaces, in our experiment the attenuated C strain induced a significantly higher production of this immunoglobulin class in the serum very early (on day 4) following vaccination. This could be ascribed to the affinity of IgA antibodies to neutralize or agglutinate virus particles. Early appearance (4 and 10 days after the vaccination) of a significantly higher concentration of IgG and IgM could be induced by the ORF virus strain D1701 applied as an adjuvant.
Thermal imaging has been utilised, both preclinically and clinically, as a tool for assessing inf... more Thermal imaging has been utilised, both preclinically and clinically, as a tool for assessing inflammation. Psoriasis is a chronic inflammatory skin disease characterised by hyperkeratosis, dermal inflammatory infiltrate and increased angiogenesis. The aim of the present study was to assess the usefulness of thermography in psoriatic lesion regression after topical treatment with bee propolis, recognised as potent antioxidants and anti-inflammatory agents. We monitored the inflammation process induced by irritants such as n-Hexyl salycilate (HXS) or Di-n-Propyl Disulfide (PPD) by histopatological assessment of the skin, thermographic scanning, total number of inflammatory cells in the peritoneal cavity, differential analysis of cells in the peritoneal cavity, macrophage spreading index, haematological and biochemical parameters, frequencies of micronucleated reticulocytes, lipid peroxidation and glutathione assay in the skin. Topically applied ethanolic extract of propolis (EEP) with HXS or PPD reduced the lipid peroxidation in the skin and total number of inflammatory cells in the skin and peritoneal cavity, functional activity of macrophages, the number of micronuclei in mouse peripheral blood reticulocytes and enzymatic activity of ALP and AST. These results demonstrate that topical application of EEP may improve psoriatic-like skin lesions by suppressing functional activity of macrophages and ROS production. Taken together, it is suggested that EEP can safely be utilised in the prevention of psoriasis-related inflammatory changes without causing any toxic effect.
In this paper we report for the first time a method for the extraction of the protein fraction fr... more In this paper we report for the first time a method for the extraction of the protein fraction from Feijoa sellowiana Berg. fruit and its electrophoretic analysis. In addition we analyzed the protein fraction for its antioxidant activity and its effectiveness against different Gram-positive and Gram-negative bacteria both as American type culture collection (ATCC) standard and clinically isolated strains. Both antimicrobial and antioxidant activity resulted stronger respect to that previously obtained for the acetonic extract from the same fruit. Further, we study both activities also on the product of in vitro gastrointestinal digestion of F. sellowiana fruit proteins. The results showed that in vitro gastrointestinal digestion increased 10 fold the antioxidant activity, while the antimicrobial activity, tested only on ATCC strains, resulted from 2 to 4 fold increased. MTT assays showed the nontoxicity of these proteins both before and after digestion.
This study was carried out to find whether propolis and its polyphenolic/flavonoids compounds may... more This study was carried out to find whether propolis and its polyphenolic/flavonoids compounds may induce cytotoxicity in primary culture of human urinary bladder transitional cell carcinoma (TCC) cells as compared to normal urinary bladder epithelial cells. Pieces of TCC or normal epithelial tissue were collected by transurethral surgery from patients in different stages (grade G1,G2, G3) of TCC. Incubation of TCC cells for cytotoxicity testing were carried out with or without diffrent concentration (50, 150, 300 µg/ml) of test components. The cytotoxicity of two preparation of propolis (water and ethanolic extract of propolis; WSDP or EEP) and its polyphenolic compounds (caffeic acid, naringin, chrysin, quercetin) was determined using trypan blue exlusion assay. Findings suggest that EEP is the most effective in inhibition of urinary bladder TCC cell proliferation as compared to WSDP or single flavonoids derived from propolis. All test components showed no cytotoxicity to normal epithelial cells. The result of this study may provide great impact on the potential activity of EEP as an adjuvant to surgery, to suppress or prevent tumor recurrence in urinary bladder since only a few anti-cancer drugs have been effective in tumor control. Since immunomodulation by BCG has been used to impruve the results of surgery it is likely that propolis preparation (EEP) as immunomodulating compound may be a substitute for mycobacterial treatment since propolis preparation or its polyphenolic components have expressed no side effect after treatment in animal models.
The aim of this study was to evaluate the in vitro influence of mitogens on the lymphoproliferati... more The aim of this study was to evaluate the in vitro influence of mitogens on the lymphoproliferative response after vaccination against classical swine fever (CSF). We also compared the effect of the ORF virus strain D1701, whose immunostimulatory effects in vivo are known, as well as the CSF virus C strain on lymphocyte cell cultures of vaccinated and control pigs. Peripheral blood leucocytes (PBL) were used in a lymphocyte proliferation assay by Concanavalin A, phytohemagglutinin, pokeweed mitogen, the attenuated ORF virus strain D1701 and the CSF virus China (C) strain. PBL were obtained from pigs vaccinated with different CSF vaccines (a subunit gp E2 vaccine, a combination of a subunit vaccine and an ORF virus strain D1701 and attenuated C strain vaccine) and from non-vaccinated pigs. Blood samples for the mitogenic response assay were collected before (day 0), and on the 4th, 10th, 21st and 28th post-vaccination day. PBL of pigs that received the C strain and ORF virus showed in vitro a weaker proliferative response to the mitogens than PBL of pigs vaccinated with the subunit CSF vaccine. The lymphocyte proliferation was observed only in pigs vaccinated with the C strain vaccine after exposure to the same strain in vitro. However, the ORF virus, in the applied concentration did not induce a significant reaction of lymphocytes that originated from either vaccinated or control animals. The results of our study suggest that the applied mitogens may induce in vitro reactivity which could play a significant immunological role after CSF vaccination. In contrary, the attenuated virus strains used as stimulators, induced no significant lymphoproliferation.
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
Using a Swiss albino mouse model of Ehrlich ascites tumor (EAT), we investigated antitumor effica... more Using a Swiss albino mouse model of Ehrlich ascites tumor (EAT), we investigated antitumor efficacy of two preparations of propolis (water and ethanolic extract) or quercetin, a compound of propolis, combined with chemotherapeutic drugs (Cisplatin and Doxorubicin). Egzamination was directed to suppression of tumor progression expressed as the percent increased life span. Propolis or quercetin were administrated ip to animals at doses of 50 mg kg-1 body weight for three consecutive days before tumor inoculation. Doxorubicin (20 mg kg-1) or cisplatin (10 mg kg-1) were given to mice two days after tumor cell inoculation. The results of log rank test between treated and control mice showed:WSDP, p=0.00291 ; WSDP + CIS, p=0.00781 ; WSDP + DOX, p=0.4998 ; EEP, p=0.00902 ; EEP + CIS, p=0.27603 ; EEP + DOX, p=0.22014 ; QU, p=0.27454 ; QU + CIS, p=0.72713 ; QU + DOX, p= 0.02667 ; CIS, p=0.00350 ; DOX, p= 0.2593. Combined treatment with test compounds and Cisplatin resulted in pozitive outcome (p=0, 00853 ; p=0, 00767 ; p=0, 02500) versus cisplatin alone, however combined treatment with doxorubicin was ineffective. Our findings indicate that WSDP, EEP, and quercetin significantly enhanced the antiproliferative and cytotoxic action of cisplatin and that propolis compounds may be usefully used for lowering the toxicity and other deteriorating activities chemotherapeutic drugs.
Kolibaciloza je glavni uzrok oboljevanja i mortaliteta odbijene prasadi. Najcesce je posljedica e... more Kolibaciloza je glavni uzrok oboljevanja i mortaliteta odbijene prasadi. Najcesce je posljedica enterotoksigenih sojeva bakterije E. coli (ETEC) koji proizvode enterotoksine sto ostecuju stijenku crijeva, poticu lucenje sluzi i elektrolita i tako uzrokuju dijareju. Vakcinacija protiv kolibaciloze odbijene prasadi nije u siroj uporabi kao profilakticka mjera. Naime, pokusaji da se potakne specificni imunosni odgovor protiv ETEC sojeva na mukoznim povrsinama crijeva nakon oralne imunizacije s fimbrijskim vakcinama bili su neucinkoviti. Zastita od mikrobnih antigena, kao sto su ETEC sojevi, na mukoznim povrsinama mogla bi se postici indukcijom i regulacijom specificnih obrambenih mehanizama posredovanim imunosnim stanicama u limfatickom tkivu probavnog sustava (LTPS). Vrednovali smo imunogenost F4ac^+ ne-ETEC soja kao izvora replicirajuceg antigena u specificnoj imunizaciji odbijene prasadi temeljem kvalitativnih i kvantitativnih pokazatelja aktivacije njihovih imunosnih stanica u mezenterijskom limfnom cvoru (MLC) uporabom protocne citometrije i imunohistologije, kao i in vitro određivanjem fagocitnih funkcija peritonejskih makrofaga. Sedam dana nakon intragastricke imunizacije sa 10^10 CFU/ml F4ac^+ ne-ETEC soja, analizirali smo CD^+/SWC^+ imunofenotipove stanica iz MLC uporabom monoklonskih protutijela specificnih za CD3, CD8, CD4, CD21, CD14, SWC3 i MHC II povrsinske stanicne molekule. U MLC imunizirane prasadi utvrdili smo porast udjela CD4^+CD8^+ memorijskih stanica, aktiviranih CD25 T pomocnickih i T citotoksicnih limfocita, B limfocita i stanica koje prikazuju antigen. Peritonejski makrofagi imunizirane prasadi imali su vecu sposobnost ingestije cestica. Izrazitija razlika u brojnosti aktiviranih CD25^+ T-limfocita vidljiva je u MLC imunizirane u odnosu na kontrolnu prasad. Prema ovim rezultatima, pokusni vakcinalni F4ac^+ ne-ETEC soj uporabljen za poticanje specificne imunosti odbijene prasadi protiv kolibaciloze izazvao je odgovore mijeloidnih i limfoidnih stanica u odjeljku LTPS tj. u MLC, i unutar peritonejske supljine odbijene prasadi.
This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY
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