Jelica Stojanovic

Immune Complexes and Complement in Serum and Synovial Fluid of Rheumatoid Arthritis Patients

Immune Complexes and Complement in Serum and Synovial Fluid of Rheumatoid Arthritis Patients Rheumatoid arthritis (RA) is predominantly an intraarticular inflammatory and autoimmune disease that involves different autoantibodies and effector mechanisms. The aim of the study was to determine the utility of Circulating Immune Complexes (CIC) and complement components (C3c, C4) as possible markers for the disease activity in laboratory diagnostics. In a cross-section study 59 patients, according to the clinical criteria, were categorized into two groups: group with moderate (MA, n=24), and group with severe activity (SA, n=35) of RA. The concentration of CIC, C3c and C4 in sera (S) and synovial fluids (SF) was examined by an immunonephelometric method in both groups and compared with values in the control group (n=15) of patients with lesions of the menisci. Obtained results showed that there was no statistical significance in the values of C3c and C4, in both biological fluids, among all tested groups. Significant differences were found in the levels of CIC in both fluids, while testing the parameters (× ± SD, IU/mL) in the sera of groups with SA and MA of RA: 7.43 ± 13.40; 3.01 ± 2.92 (p<0.05) and SF: 13.47 ± 21.1, 5.33 ± 7.53 (p<0.001), respectively. These differences were higher between the group with SA and CG. Results for the concentrations of CIC were significantly higher in SF compared to sera: in the RA group with SA by 77% and group with MA by about 82%. These data could provide a confirmation of the hypothesis about local, intraarticular autoantibodies and subsequent CIC production. It can be concluded that the examination of CIC concentration in serum, and where it is possible in SF, is a useful marker of disease activity in RA patients, in contrast to the tested components of the complement. This statement does not exclude their consumption within immune effector mechanisms, but elicits the possibility that lower molecular fragments (C3d, C4d), as well as the novel activation products, could be better disease activity markers in RA patients. Imunski Kompleksi i Komplement u Serumu i Sinovijalnoj Tečnosti Kod Bolesnika sa Reumatoidnim Artritisom Reumatoidni artritis (RA) jeste predominantno intraartikularna zapaljenska i autoimunska bolest u koju su uključena različita autoantitela i efektorni mehanizmi. Cilj ispitivanja je bio da se ustanovi značaj cirkulišućih imunskih kompleksa (CIK) i komponenti komplementa (C3c, C4), kao pokazatelja stepena aktivnosti RA za laboratorijsku dijagnostiku. U studiji preseka stanja je ispitano 59 bolesnika koji su prema kliničkim kriterijumima za aktivnost RA podeljeni u dve grupe: grupu sa umerenom (UA, n=24) i grupu sa visokom aktivnošću (VA, n=35) RA. Koncentracije CIK, C3c i C4 u serumu i sinovijalnoj tečnosti (ST) određivane su imunonefelometrijskom metodom u obe grupe ispitanika i upoređene sa vrednostima u kontrolnoj grupi od 15 pacijenata s povredama meniskusa. Rezultati su pokazali da nije bilo statistički značajnih razlika u koncentracijama za C3c i C4 u oba biološka uzorka između ispitivanih grupa. Statistički značajne razlike u koncentracijama CIK utvrđene testiranjem vrednosti (× ± SD, IU/mL) u serumu između grupe sa VA i grupe sa UA RA bile su: 7,43 ± 13,40; 3,01 ± 2,92 (p < 0,05) i za vrednosti u ST: 13,47 ± 21,1, 5,33 ± 7,53 (p < 0,001). Razlike su bile više izražene između grupe sa VA RA i KG. Rezultati koncentracija CIK su bili značajno viši u ST u odnosu na serum u obe grupe bolesnika: u grupi sa UA za 77% a u grupi sa VA RA za 82%. Ti podaci idu u prilog potvrdi hipoteze o lokalnoj, intraartikularnoj produkciji autoantitela, odnosno CIK. Može se zaključiti da je laboratorijsko određivanje koncentracije CIK korisan pokazatelj stepena aktivnosti RA, što se ne odnosi na ispitivane komponente komplementa. To ne isključuje njihovu aktivnost u okviru efektornog imunskog mehanizma, ali ukazuje na to da bi manji molekulski fragmenti (C3d, C4d) i novi aktivacioni produkti mogli biti bolji pokazatelji stepena aktivnosti RA.

Detergent-like stressor and nutrient in metabolism of Penicillium chrysogenum

The influence of detergents on the metabolism of Penicillium chrysogenum from two aspects, as a stress factor and potential nutrient, was studied. The fungus was isolated from the river bed Lepenica, Kragujevac, at a place where sewage domestic wastewater discharged into the river. The fungus was grown in a liquid nutrient medium according to Czapek with and without addition of commercial detergent (MERIX, Henkel, Serbia) at a concentration of 0.3% and 0.5%. The biochemical changes of pH, redox potential, free and total organic acids, total dry weight biomass, activity of alkaline and acid invertase and alkaline phosphatase were evaluated from day 3 to day 16 of the fungus growth. At the same time, detergent disappearance in terms of methylene blue active substances in the medium was measured. The detergent at a concentration of 0.5% showed a fungicide effect. In the medium with 0.3% of detergent, there was increased pH and concentration of organic acids, but decreased redox potential and total dry weight biomass. The detergent also showed an inhibitory effect on invertase and phosphatase activity. P. chrysogenum decomposed 50.2% of the total detergent concentration for an experimental period of 16 days.

Urinary Cystatin C as a Marker of Tubular Dysfunction

Urinary Cystatin C as a Marker of Tubular Dysfunction Cystatin C (CysC) is a nonglycosylated 13 KD protein that belongs to the type II cystatin gene family. It is a strong inhibitor of cysteine proteinases, freely filtered by the kidney glomerulus and reabsorbed by the tubulus, where it is almost totally catabolized. Remainder of the nonmetabolized CysC is eliminated in urine and may represent a useful marker of tubular dysfunction. The aim of the study was to confirm the clinical importance of the quantitative determination of CysC by an automated immunonephelometric method (DADE Behring). Two groups of patients were examined: one with glomerular (GD, n=36) and one with tubular dysfunction (TD, n=31), and compared with the control group (CG, n=31) of healthy males and females from laboratory personnel (n=11) and patients on routine systematic examination (n=20), from 25 to 58 years old. The patient groups were categorised according to the urine analysis of total proteins, creatinine and adequate proteins electrophoretic panel. CysC concentration in CG was in the range of 0.02-0.15 mg/L; 0.01-0.48 mg/L and 0.25-18 mg/L in GD and TD groups respectively. Values of means ± SD for patient groups (GD=0.11 ± 0.14; TD=3.92 ± 3.75 mg/L) showed statistical significance (p<0.001) in the TD group in relation to GD and CG groups. It confirms that quantitative determination of CysC in one urine portion, with a fast laboratory method, might be a useful marker of tubular dysfunction, especially in emergency situations, taking into account that there is no interference of circadian variation on its concentration. Cistatin C U Urinu Kao Pokazatelj Tubulskog Oštećenja Cistatin C (CysC) je neglikozirajući protein molekulske mase od 13 KD koji pripada tipu II genske familije cistatina. Ima funkciju inhibitora cistein proteinaze koji se potpuno filtrira u glomerulima a zatim reapsorbuje i kataboliše u proksimalnim tubulima. Preostali i nerazgrađen CysC biva eliminisan urinom pa zato može da bude koristan marker tubulskih oštećenja. Cilj ispitivanja bio je da se utvrdi klinički značaj kvantitativnog određivanja CysC automatizovanom imunonefelometrijskom metodom (DADE Behring). Ispitane su dve grupe bolesnika: sa tubulskom (TI) i glomerulskom (GI) insuficijencijom, i upoređene sa kontrolnom grupom (KG) koju su činile zdrave osobe oba pola (n=31), laboratorijski personal (n=11) i pacijenti na sistematskom pregledu (n = 20), starosne dobi od 25 do 58 godina. Kategorizacija grupa sa GI (n=36) i TI (n=31) ustanovljena je na osnovu koncentracije ukupnih proteina, kreatinina i adekvatnih elektroforetskih denzitometrijskih varijanti karakterističnih za obe grupe. Koncentracija CysC u KG bila je u opsegu 0,02-0,15 mg/L, u grupi sa GI 0,01-0,48 mg/L, a u grupi sa TI 0,25-18 mg/L. Rezultati vrednosti (medijana±SD) za grupu sa GI (0,11 ± 0,14 mg/L) i grupu sa TI (3,92 ± 3,75 mg/L) pokazali su statistički značajnu razliku (p< 0,0001) između grupe sa TI i grupa sa GI i KG. Pokazano je da određivanje CysC u jednom uzorku urina brzim laboratorijskim postupkom može biti koristan test za diferencijalnu dijagnostiku tubulskih u odnosu na glomerulska oštećenja, posebno u urgentnim stanjima, s obzirom na činjenicu da na njegovu koncentraciju nema uticaja cirkadijalnih ritmova.

Multiple myeloma invasion of the central nervous system

INTRODUCTION Multiple myeloma (MM) is characterized by the presence of neoplastic proliferating plasma cells. The tumor is generally restricted to the bone marrow. The most common complications include renal insufficiency, hypercalcemia, anemia and reccurent infections. The spectrum of MM neurological complications is diverse, however, involvement of MM in the cerebrospinal fluid (CSF) and leptomeningeal infiltration are rare considered. In about 1% of the cases, the disease affects the central nervous system (CNS) and presents itself in the form of localized intraparenchymal lesions, solitary cerebral plasmocytoma or CNS myelomatosis (LMM). CASE REPORT We presented the clinical course of a 55-year-old man with MM and LMM proven by malignant plasma cells in the CSF, hospitalized with the pain in the thoracic spine. His medical history was uneventful. There had been no evidence of mental or neurological impairment prior to the seizures. Physical examination showed no abnormalities. After a complete staging, the diagnosis of MM type biclonal gammopathia IgG lambda and free lambda light chains in the stage III was confirmed. The treatment started with systemic chemotherapy (with vincristine, doxorubicin plus high-dose dexamethasone--VAD protocol), radiotherapy and bisphosphonate. The patient developed weakness, nausea, febrility, dispnea, bilateral bronchopneumonia, acute renal insufficiency, confusions, headaches and soon thereafter sensomotor aphasias and right hemiparesis. The patient was treated with the adequate therapy including one hemodyalisis. His neurological status was deteriorated, so Multislice Computed Tomography (MSCT) of the head was performed and the findings were normal. Analysis of CSF showed pleocytosis, 26 elements/mL and increased concentrations of proteins. Cytological analysis revealed an increased number of plasma cells (29%). Electrophoretic analysis of proteins disclosed the existance of monoclonal components in the serum, urine and CSF. Immunofixation electrophoretic and quantitative nephelometric tests confirmed Biclonal multiple myeloma of IgG lambda and light chain lambda isotypes. Analysis of neurothropic viruses with ELISA methods was negative. Once the presence of LMM was confirmed, the patient received intrathecal chemotherapy with methotrexate, cytosine arabinoside, dexamethasone three times a week, and systemic high doses of dexamethasone iv like a single agent without craniospinale irradiations. Despite the treatment, the patient died one month after the diagnosis. Autopsy was not performed. CONCLUSION Presented patient, as well as most other patients with MM progressing to CN Sinfiltration was in the stage III. In addition to the detailed clinical examination, and all investigations required for MM diagnosis and staging of the disease, we introduced the additional CSF examination and calculation of kappa lambda ratio, that helped us make an early diagnosis and prognosis of MM with LMM. Although LMM had a low prevalence, it could be more frequent than expected especially in patients with high risk. CSF examination with positive plasma cells and abnormal morphology remains the hallmark for diag nosing CNS infiltration.

Free Light Chains of Immunoglobulin as a Prognostic Factor for Some Plasmaproliferative Diseases

Free Light Chains of Immunoglobulin as a Prognostic Factor for Some Plasmaproliferative Diseases Quantitation of monoclonal immunoglobulins and their fragments is used for monitoring the plasmaproliferative disease course and the effect of therapy. The aim of free light chains examination was to evaluate the significance of the FLC ratio as a prognostic factor for remission, progression and survival in different disease groups. The concentrations of immunoglobulins and free light chains were measured by an immunonephelometric method on a »SIEMENS« DADE BN II analyser with reagents (Freelite, The Binding Site, UK). In this examination 151 patients from 3 different disease groups: 1. Light chain disease or Bence Jones myeloma (37), 2. Biclonal gammopathy with FLC (23) and 3. Monoclonal gammopathy of undetermined significance (91), were investigated during a period of 7 years. The reference interval for FLC ratio is 0.26-1.65. According to the International Staging System for multiple myeloma, a serum FLC ratio of <0.03 or >32 was taken as abnormal. The patients with light chain disease and biclonal gammopathy with FLC with an abnormal FLC ratio and a combination of adverse risk factors (76.7%) had median survival times of 22-30 months, versus patients with a normal or slightly varied FLC ratio without adverse risk factors (23.3%) with median survival times of 39-51 months. About 38% of patients who had shown lowered free light chains values by more than 50% under therapy, achieved disease remission in the light chain disease and biclonal gammopathy with FLC groups. In the group of patients with monoclonal gammopathy of undetermined significance, 66.0% had a normal or slightly modified FLC ratio which corresponds to low and low-intermediate risk of disease progression, as opposed to 34.0% with an abnormal FLC ratio (<0.25 or >4) which corresponds to high and high-intermediate risk. An abnormal FLC ratio in the examined groups could be an independent risk factor for progression and poorer disease prognosis. Slobodni Laki Lanci Imunoglobulina Kao Prognostički Faktor Kod Nekih Plazmaproliferativnih Bolesti Kvantitativno određivanje monoklonskih imunoglobulina i njihovih fragmenata koristi se za praćenje toka i terapijskog odgovora kod plazmaproliferativnih bolesti. Cilj određivanja slobodnih lakih lanaca imunoglobulina u serumu bolesnika jeste provera značaja njihovog količnika (κ/λ indeks) kao prognostičkog faktora remisije, progresije i preživljavanja. Koncentracije imunoglobulina i slobodnih lakih lanaca određivane su imunonefelometrijskom metodom na analizatoru SIEMENS DADE Behring II sa reagensima (FREELITE, The Binding Site, UK). U ispitivanje je uključen 151 bolesnik tokom perioda od 7 godina, koji su razvrstani u 3 grupe: 1. bolest lakih lanaca ili Bence Jones mijelom (37); 2. biklonalna gamapatija sa slobodnim lakim lancima (23) i 3. monoklonska gamapatija neutvrđenog značaja (91). Referentnim intervalom za κ/λ indeks smatraju se vrednosti 0,26-1,65. Prema Internacionalnom prognoznom indeksu za multipli mijelom, kao patološki uzet je κ/λ indeks <0,03 ili >32. Bolesnici iz prve dve grupe sa patološkim k/λ indeksom i kombinacijom nepovoljnih faktora rizika (76,7%) imali su prosečno vreme preživljavanja 22-30 meseci, nasuprot bolesnicima sa fiziološkim ili neznatno izmenjenim κ/λ indeksom bez nepovoljnih faktora rizika (23,3%), sa prosečnim vremenom preživljavanja 39-51 mesec. Oko 38% bolesnika koji su pod terapijom imali sniženje κ/λ indeksa >50% su ostvarili remisiju bolesti. U grupi ispitanika sa MGNZ, 66,0% je imalo fiziološke ili neznatno izmenjene κ/λ indekse, što odgovara niskom i srednje niskom riziku progresije, nasuprot 34,0% sa patološkim κ/λ indeksom (<0,25 ili >4), što odgovara srednje visokom i visokom riziku progresije. Postojanje patološki značajnog κ/λ indeksa u ispitivanim grupama predstavlja nezavisan faktor rizika za progresiju bolesti i lošiju prognozu.

The spectral analysis of motion: An "open field" activity test example

In this work we have described the new mathematical approach, with spectral analysis of the data to evaluate position and motion in the „„open field““ experiments. The aim of this work is to introduce several new parameters mathematically derived from experimental data by means of spectral analysis, and to quantitatively estimate the quality of the motion. Two original software packages (TRACKER and POSTPROC) were used for transforming a video data to a log file, suitable for further computational analysis, and to perform analysis from the log file. As an example, results obtained from the experiments with Wistar rats in the „open field“ test are included. The test group of animals was treated with diazepam. Our results demonstrate that all the calculated parameters, such as movement variability, acceleration and deceleration, were significantly lower in the test group compared to the control group. We believe that the application of parameters obtained by spectral analysis could be of great significance in assessing the locomotion impairment in any kind of motion. [Projekat Ministarstva nauke Republike Srbije, br. III41007 i br. ON174028]

ANALYSIS OF THE AMINO ACIDS OF SOME TYPES OF FUNGI CULTIVATED IN THE PRESENCE OF DETERGENT

The paper presents the results of the influence of anionic-type detergent containing sodium tripolyphosphate and ethoxylated oleylcetyl alcohol on the type and quantity of present amino acids in the fermentation broths of the fungi Aspergillus niger, Alternaria tenuis and Fusarium oxysporum cultivated on a medium with the addition of 1% detergent as a function of their application in the process of detergent biodegradation. In the case of A.niger the stimulation of the biosynthesis of 15 various amino acids was determined. In the case of A.tenuis the production of 14 amino acids was inhibited as compared to the total number of 15 amino acids identified when the fungi were cultivated in the absence of detergent, whereby the explicit inhibition of the synthesis of methionine, isoleucine and leucine was registered. Detergent (1% concentration) exhibits a stimulating effect on the bioproduction of amino acids in the case of the fungus F.oxysporum when the presence of 14 amino acids was identified.

EFFECTS OF 3-METHYLHISTAMINE AND PHENYLETHYLAMINE ON HISTAMINE ACTION ON ISOLATED GUINEA-PIG TRACHEA RINGS

It is well known that histamine produces constriction via H1 receptors and decreases tracheal smooth muscle tone via H2 and H3 receptors. In addition, it has already been reported that 3-methylhistamine and phenylethylamine are competitive antagonists of histamine N-methyl-transferase (HMT), the enzyme responsible for rapid inactivation of histamine. Our results suggest the possibility that 3- methyl-histamine and phenylethylamine as competitive antagonists of histamine N-methyl-transferase lead to potentiation of histamine induced constriction of isiolated guinea-pig trachea, probably due to the decrease of histamine methylation and consequent inactivation. In as much, 3-methyl-histamine and phenylethylamine had no effect on the basal tone of isolated trachea smooth muscle, as well as on other mechanisms leading to increased responsiveness of guinea-pig tracheal smooth muscle (acetylcholine, KCl, electro stimulation).