Infection of neurons with cytomegalovirus

Infekcija HCMV-om, pripadnikom porodice β herpesvirusa, široko je rasprostranjena u ljudskoj populaciji. Kod imunokompetentnih pojedinaca infekcija uzrokuje blaže ili nikakve simptome, dok kod osoba sa oslabljenim ili nezrelim imunološkim sustavom može biti životno ugrožavajuća. Kao najčešći uzročnik kongenitalnih virusnih infekcija, HCMV uzrokuje teška strukturalna i funkcionalna oštećenja CNS-a. Trenutno ne postoji cjepivo za citomegalovirus, a dostupni terapijski postupci pokazuju ograničenu učinkovitost, stoga su istraživanja temeljnih mehanizama nastanka bolesti od izuzetne važnosti. Skoro sva dostupna istraživanja infekcije neurona CMV-om provedena su u in vitro uvjetima i pokazuju oprečne rezultate o sklonosti CMV-a prema neuronima. Svrha ovog rada bila je provesti in vivo istraživanje na mišjem modelu s reporterskim MCMV virusom kako bi se otkrilo mogu li neuroni biti produktivno inficirani. Rezultati su pokazali da u mozgu miševa dolazi do in vivo konverzije reporterskog virusa 14. dan nakon infekcije. Ovi rezultati ukazuju da inficirani neuroni produciraju virus, ali zbog niskog titra reporterskog virusa u mozgu korištenih miševa nismo saznali koliki postotak neurona ima sposobnost produkcije virusnih čestica. Stoga, ovo istraživanje služi kao preliminarno za daljnja in vivo istraživanja citomegalovirusne infekcije neurona.Infection with HCMV, a member of the β herpesvirus family, is widely distributed within the human population. In immunocompetent individuals the infection causes mild or none symptoms. On the other hand, it can be a life-threatening infection in people with weakened or immature immune system. As the most common cause of congenital viral infections, HCMV leads to severe structural and functional abnormalities of the CNS. At the moment there is no vaccine to prevent cytomegalovirus infection. Also, the available therapeutic procedures show limited efficacy. Therefore, the research on basic mechanisms of CMV infection are of great importance. Almost all the available research on CMV infection of neurons have been conducted in vitro and suggest conflicting results on CMV preference for neurons. The purpose of this paper was to conduct an in vivo research using mouse model with reporter MCMV virus to determine whether neurons can be productively infected. The results have demonstrated that in vivo conversion of the reporter virus occurs in the brain of mice 14 days after the infection. These results indicate that infected neurons can produce the virus. However, due to the low titer of reporter virus in the brain of mice, we haven't found out the exact percentage of neurons that has the ability to produce the virus. In conclusion, this paper serves as a preliminary research for necessary further in vivo research on CMV infection of neurons

Infection of neurons with cytomegalovirus

Infekcija HCMV-om, pripadnikom porodice β herpesvirusa, široko je rasprostranjena u ljudskoj populaciji. Kod imunokompetentnih pojedinaca infekcija uzrokuje blaže ili nikakve simptome, dok kod osoba sa oslabljenim ili nezrelim imunološkim sustavom može biti životno ugrožavajuća. Kao najčešći uzročnik kongenitalnih virusnih infekcija, HCMV uzrokuje teška strukturalna i funkcionalna oštećenja CNS-a. Trenutno ne postoji cjepivo za citomegalovirus, a dostupni terapijski postupci pokazuju ograničenu učinkovitost, stoga su istraživanja temeljnih mehanizama nastanka bolesti od izuzetne važnosti. Skoro sva dostupna istraživanja infekcije neurona CMV-om provedena su u in vitro uvjetima i pokazuju oprečne rezultate o sklonosti CMV-a prema neuronima. Svrha ovog rada bila je provesti in vivo istraživanje na mišjem modelu s reporterskim MCMV virusom kako bi se otkrilo mogu li neuroni biti produktivno inficirani. Rezultati su pokazali da u mozgu miševa dolazi do in vivo konverzije reporterskog virusa 14. dan nakon infekcije. Ovi rezultati ukazuju da inficirani neuroni produciraju virus, ali zbog niskog titra reporterskog virusa u mozgu korištenih miševa nismo saznali koliki postotak neurona ima sposobnost produkcije virusnih čestica. Stoga, ovo istraživanje služi kao preliminarno za daljnja in vivo istraživanja citomegalovirusne infekcije neurona.Infection with HCMV, a member of the β herpesvirus family, is widely distributed within the human population. In immunocompetent individuals the infection causes mild or none symptoms. On the other hand, it can be a life-threatening infection in people with weakened or immature immune system. As the most common cause of congenital viral infections, HCMV leads to severe structural and functional abnormalities of the CNS. At the moment there is no vaccine to prevent cytomegalovirus infection. Also, the available therapeutic procedures show limited efficacy. Therefore, the research on basic mechanisms of CMV infection are of great importance. Almost all the available research on CMV infection of neurons have been conducted in vitro and suggest conflicting results on CMV preference for neurons. The purpose of this paper was to conduct an in vivo research using mouse model with reporter MCMV virus to determine whether neurons can be productively infected. The results have demonstrated that in vivo conversion of the reporter virus occurs in the brain of mice 14 days after the infection. These results indicate that infected neurons can produce the virus. However, due to the low titer of reporter virus in the brain of mice, we haven't found out the exact percentage of neurons that has the ability to produce the virus. In conclusion, this paper serves as a preliminary research for necessary further in vivo research on CMV infection of neurons

Dissecting the cytomegalovirus CC chemokine: Chemokine activity and gHgLchemokine-dependent cell tropism are independent players in CMV infection

Like all herpesviruses, cytomegaloviruses (CMVs) code for many immunomodulatory proteins including chemokines. The human cytomegalovirus (HCMV) CC chemokine pUL128 has a dual role in the infection cycle. On one hand, it forms the pentameric receptor-binding complex gHgLpUL(128,130,131A), which is crucial for the broad cell tropism of HCMV. On the other hand, it is an active chemokine that attracts leukocytes and shapes their activation. All animal CMVs studied so far have functionally homologous CC chemokines. In murine cytomegalovirus (MCMV), the CC chemokine is encoded by the m131/m129 reading frames. The MCMV CC chemokine is called MCK2 and forms a trimeric gHgLMCK2 entry complex. Here, we have generated MCK2 mutant viruses either unable to form gHgLMCK2 complexes, lacking the chemokine function or lacking both functions. By using these viruses, we could demonstrate that gHgLMCK2-dependent entry and MCK2 chemokine activity are independent functions of MCK2 in vitro and in vivo. The gHgLMCK2 complex promotes the tropism for leukocytes like macrophages and dendritic cells and secures high titers in salivary glands in MCMV-infected mice independent of the chemokine activity of MCK2. In contrast, reduced early antiviral T cell responses in MCMV-infected mice are dependent on MCK2 being an active chemokine and do not require the formation of gHgLMCK2 complexes. High levels of CCL2 and IFN-gamma in spleens of infected mice and MCMV virulence depend on both, the formation of gHgLMCK2 complexes and the MCK2 chemokine activity. Thus, independent and concerted functions of MCK2 serving as chemokine and part of a gHgL entry complex shape antiviral immunity and virus dissemination. Studies on immunomodulatory proteins of herpesviruses have strongly contributed to understanding antiviral immune responses elicited during infection and to defining targets for intervention during herpesvirus infections. CC chemokines of cytomegaloviruses have been shown to shape the viral cell tropism and antiviral immune responses, yet, their modes of action are not really understood. Here, we used well-defined mutants of the MCMV chemokine MCK2 to exemplarily dissect the role of a CMV CC chemokine in viral dissemination and antiviral innate and adaptive immune responses. We could show that the chemokine activity and the entry function of MCK2 are independent players in the MCMV infection of the mouse. As cytomegaloviruses are discussed as potential vaccine vectors which may be programmed to elicit specific CD8+ T cell responses and combat specific pathogens, it is important to understand the contribution of immunomodulatory proteins to vector-induced immune responses. Specifically, in infections of Rhesus macaques with Rhesus CMV (RhCMV), the viral CC chemokine has been shown to drastically shape the immune response to vaccine antigens. Our detailed analysis of the dual roles of a CMV CC chemokine may further help to decide whether CMV vaccine vectors should express an intact viral CC chemokine or not

Fast, Reliable, and Simple Point-of-Care-like Adaptation of RT-qPCR for the Detection of SARS-CoV-2 for Use in Hospital Emergency Departments

During COVID-19 pandemics, the availability of testing has often been a limiting factor during patient admissions into the hospital. To circumvent this problem, we adapted an existing diagnostic assay, Seegene Allplex SARS-CoV-2, into a point-of-care-style direct qPCR (POC dqPCR) assay and implemented it in the Emergency Department of Clinical Hospital Center Rijeka, Croatia. In a 4-month analysis, we tested over 10, 000 patients and demonstrated that POC-dqPCR is robust and reliable and can be successfully implemented in emergency departments and similar near-patient settings and can be performed by medical personnel with little prior experience in qPCR

Memory CD8 T Cells Generated by Cytomegalovirus Vaccine Vector Expressing NKG2D Ligand Have Effector-Like Phenotype and Distinct Functional Features

Viral vectors have emerged as a promising alternative to classical vaccines due to their great potential for induction of a potent cellular and humoral immunity. Cytomegalovirus (CMV) is an attractive vaccine vector due to its large genome with many non-essential immunoregulatory genes that can be easily manipulated to modify the immune response. CMV generates a strong antigen-specific CD8 T cell response with a gradual accumulation of these cells in the process called memory inflation. In our previous work, we have constructed a mouse CMV vector expressing NKG2D ligand RAE-1γ in place of its viral inhibitor m152 (RAE-1γMCMV), which proved to be highly attenuated . Despite attenuation, RAE-1γMCMV induced a substantially stronger CD8 T cell response to vectored antigen than the control vector and provided superior protection against bacterial and tumor challenge. In the present study, we confirmed the enhanced protective capacity of RAE-1γMCMV as a tumor vaccine vector and determined the phenotypical and functional characteristics of memory CD8 T cells induced by the RAE-1γ expressing MCMV. RNAseq data revealed higher transcription of numerous genes associated with effector-like CD8 T cell phenotype in RAE-1γMCMV immunized mice. CD8 T cells primed with RAE-1γMCMV were enriched in TCF1 negative population, with higher expression of KLRG1 and lower expression of CD127, CD27, and Eomes. These phenotypical differences were associated with distinct functional features as cells primed with RAE-1γMCMV showed inferior cytokine-producing abilities but comparable cytotoxic potential. After adoptive transfer into naive hosts, OT-1 cells induced with both RAE-1γMCMV and the control vector were equally efficient in rejecting established tumors, suggesting the context of latent infection and cell numbers as important determinants of enhanced anti-tumor response following RAE-1γMCMV vaccination. Overall, our results shed new light on the phenotypical and functional distinctness of memory CD8 T cells induced with CMV vector expressing cellular ligand for the NKG2D receptor

ChAdOx1‐S adenoviral vector vaccine applied intranasally elicits superior mucosal immunity compared to the intramuscular route of vaccination

COVID‐19 vaccines prevent severe forms of the disease, but do not warrant complete protection against breakthrough infections. This could be due to suboptimal mucosal immunity at the site of virus entry, given that all currently approved vaccines are administered via the intramuscular route. In this study, we assessed humoral and cellular immune responses in BALB/c mice after intranasal and intramuscular immunization with adenoviral vector ChAdOx1‐S expressing full‐length Spike protein of SARS‐CoV‐2. We showed that both routes of vaccination induced a potent IgG antibody response, as well as robust neutralizing capacity, but intranasal vaccination elicited a superior IgA antibody titer in the sera and in the respiratory mucosa. Bronchoalveolar lavage from intranasally immunized mice efficiently neutralized SARS‐CoV‐2, which has not been the case in intramuscularly immunized group. Moreover, substantially higher percentages of epitope‐specific CD8 T cells exhibiting a tissue resident phenotype were found in the lungs of intranasally immunized animals. Finally, both intranasal and intramuscular vaccination with ChAdOx1‐S efficiently protected the mice after the challenge with recombinant herpesvirus expressing the Spike protein. Our results demonstrate that intranasal application of adenoviral vector ChAdOx1‐S induces superior mucosal immunity and therefore could be a promising strategy for putting the COVID‐19 pandemic under control

Slagalica nasljeđa : priručnik za opismenjavanje iz medicinske genetike

"Slagalica nasljeđa" - priručnik za opismenjavanje iz medicinske genetike ima tri namjene. Prije svega, on je edukativna slikovnica za studente, liječnike i pacijente, ali i druge zainteresirane pojedince jer su u njoj kroz ilustracije objašnjene osnove genetike čovjeka, kao i osnove medicinske genetike. Od toga kako prepoznati osobu s genetičkim poremećajem, kako nastaju i koje vrste genetičkih poremećaja postoje pa sve do toga na koji ih način možemo dijagnosticirati. Nadalje, nakon svake ilustracije na pojedinoj stranici nalaze se definicije 79 pojmova iz medicinske genetike koje čine tezaurus za studente, liječnike i pacijente koji se na bilo koji način susreću s genetičkim poremećajima. Naposljetku, ova knjiga sadrži i primjere rečenica u koje su ubačeni stručni pojmovi iz medicinske genetike, a koji su namijenjeni studentima prilikom savladavanja komunikacijskih vještina na kolegiju Medicinska genetika, ali i liječnicima prilikom informiranja svojih pacijenata o (mogućem) genetičkom poremećaju. Uz kreatoricu ideje i urednicu izdanja, doc. dr. sc. Ninu Perezu, autori izdanja su studenti šeste godine Integriranog preddiplomskog i diplomskog sveučilišnog studija Medicina i prof. dr. sc. Saša Ostojić

Slagalica nasljeđa : priručnik za opismenjavanje iz medicinske genetike

"Slagalica nasljeđa" - priručnik za opismenjavanje iz medicinske genetike ima tri namjene. Prije svega, on je edukativna slikovnica za studente, liječnike i pacijente, ali i druge zainteresirane pojedince jer su u njoj kroz ilustracije objašnjene osnove genetike čovjeka, kao i osnove medicinske genetike. Od toga kako prepoznati osobu s genetičkim poremećajem, kako nastaju i koje vrste genetičkih poremećaja postoje pa sve do toga na koji ih način možemo dijagnosticirati. Nadalje, nakon svake ilustracije na pojedinoj stranici nalaze se definicije 79 pojmova iz medicinske genetike koje čine tezaurus za studente, liječnike i pacijente koji se na bilo koji način susreću s genetičkim poremećajima. Naposljetku, ova knjiga sadrži i primjere rečenica u koje su ubačeni stručni pojmovi iz medicinske genetike, a koji su namijenjeni studentima prilikom savladavanja komunikacijskih vještina na kolegiju Medicinska genetika, ali i liječnicima prilikom informiranja svojih pacijenata o (mogućem) genetičkom poremećaju. Uz kreatoricu ideje i urednicu izdanja, doc. dr. sc. Ninu Perezu, autori izdanja su studenti šeste godine Integriranog preddiplomskog i diplomskog sveučilišnog studija Medicina i prof. dr. sc. Saša Ostojić