Examination of humoral and cellular immune responses after immunization with subunit vaccine against Herpes simplex virus 1 and 2

Abstract

The objective of our study was to examine the humoral and cellular immune response in mice after immunization with bivalent subunit vaccine against Herpes simplex viruses (HSV 1 and HSV2). These examinations were carried out in biological assay on 27 female Swiss albino mice (20 g bm). The subunit vaccine was prepared from purified glycoprotein antigens (subunits) isolated from the external envelopes of HSV 1 and HSV 2 and adsorbed on the adjuvant AI (OH)3 at a final concentration of 5 mg/ml. This vaccine contained 0.14 mg/ml total protein for the HSV 1 subunits and 0.16 mg/ml for HSV2 subunits. Two groups of 9 and 7 experimental mice were immunized with the HSV1/HSV2 subunit vaccine (0.5 ml s/c). A group of 11 healthy non-immunized mice served as the control. The specific humoral immune response of the vaccinated mice was examined by a standard method of indirect immunofluorescence. The specific cellular immune response of the vaccinated animals was tested by measuring the lymphocyte proliferation response to HSV1 and HSV2 antigens with tritiated methyl thymidine (*128.5 Bq/mmol), in vitro. The titers of the specific IgG and IgM class antibodies, in the blood sera of the first group of 9 immunized mice, on day 10 after vaccination were: from 1:8 to 1:64 IgG and 1:32 to 1:128 IgM for HSV1 and 1:32 to 1:128 IgG and IgM antibodies for HSV2. The equivalent titers for the second group of 7 immunized mice, on day 24 after vaccination were: from 1:64 to 1:128 IgG and 1:16 to 1:64 IgM for HSV1; from 1:32 to 1:128 IgG and IgM for HSV2. The mean values for radioactivity in the proliferation test of lymphocytes for HSV1 and HSV2 antigens were 1471 cpm and 1839 cpm respectively on day 10 after vaccination, The mean value for radioactivity in the medium after spontaneous proliferation of lymphocytes for this group of mice was 893 cpm. The mean radioactivity of lymphocytes from the second group of mice, stimulated with HSV1 antigens was 4158 cpm on day 24 after vaccination, with a spontaneous proliferation value of 364 cpm. Thus, the low subunit antigen concentrations in the bivalent subunit vaccine, induced satisfactory humoral and cellular immune responses to HSV 1 and HSV2 in the vaccinated mice.

Cilj naših istraživanja je bilo ispitivanje humoralnog i ćelijskog imunskog odgovora posle imunizacije miševa eksperimentalnom bivalentnom subjediničnom vakcinom protiv Herpes simplex virusa tipa 1 i 2 (HSV1 i HSV2). Ova ispitivanja su izvršena u biološkom ogledu na 27 ženki Swiss albino miševa, težine 20 g. Subjedinična vakcina je pripremljena od prečišćenih glikoproteinskih antigena (subjedinica) izolovanih iz spoljašnjih omotača HSV1 i HSV2 i adsorbovanih na adjuvans A1(OH)3. Finalna koncentracija A1(OH)3 u vakcini iznosila je 5 mg/ml. Ukupne koncentracije antigena u vakcini iznosile su 0.14 mg/ml za HSV1 i 0.16 mg/ml za HSV2. Dve grupe od 9 i 7 eksperimentalnih miševa imunizovane su supkutano sa istim dozama od 0.5 ml HSV1/HSV2 subjedinične vakcine. dok je grupa od 11 zdravih, neimunizovanih miševa predstavljala kontrolnu grupu. Specifični humoralni imunski odgovor vakcinisanih rniševa ispitivan je standardnom metodom indirektne imunofluorescencije. Specifični celularni imunski odgovor vakcinisanih životinja ispitivan je in vitro standardnom metodom proliferacije limfocita na HSV1 i HSV2 antigene pomoću tricijum metil timidina (128.5 Bq/mmol). Specifična antitela Ig G i Ig M klase ustanovljena su u krvnom serumu prve grupe miševa, desetog dana posle vakcinacije i njihov titar se kretao u opsegu od 1:8 do 1:64 (Ig G) i od 1:32 do 1:128 (Ig M) za HSV1 i od 1:32 do 1:128 (Ig G i Ig M) za HSV2. Titar specifičnih antitela u serumu miševa druge grupe, 24-tog dana posle vakcinacije, kretao se u opsegu od 1:64 do 1:128 (IgG) i 1:16 do 1:64 (Ig M) za HSV1 i od 1:32 do 1:128 (IgG i IgM) za HSV2. Srednje vrednosti testa proliferacije limfocita stimulisanih sa HSV1 i HSV2 antigenima kod prve grupe miševa, 10 dana posle imunizacije, iznosile su 1471 c/min (za HSV1) i 1839 c/min za HSV2. Srednja vrednost radioaktivnosti uzoraka u testu spontane proliferacije limfocita kod ovih miševa u medijumu, bila je 893 c/min. Srednja vrednost radioaktivnosti limfocita stimulisanih sa HSV1 antigenima u drugoj grupi miševa, 24-tog dana od vakcinacije iznosila je 4158 c/min, dok je vrednost spontane proliferacije limfocita ovih miševa bila 364 c/min. Niske koncentracije subjediničnih antigena u vakcini indukovale su zadovoljavajući humoralni i ćelijski imunski odgovor kod vakcinisanih miševa protiv HSV1 i HSV2.