Where are we with pre-exposure prophylaxis use in Central and Eastern Europe? Data from the Euroguidelines in Central and Eastern Europe (ECEE) Network Group.

OBJECTIVES: Pre-exposure prophylaxis (PrEP) for HIV infection is an important intervention for control of the HIV epidemic. The incidence of HIV infection is increasing in the countries of Central and Eastern Europe (CEE). Therefore, we investigated the change in PrEP use in CEE over time. METHODS: The Euroguidelines in Central and Eastern Europe (ECEE) Network Group was initiated in February 2016 to compare standards of care for HIV and viral hepatitis infections in CEE. Data on access to PrEP were collected from 23 countries through online surveys in May-June 2017 (76 respondents) and in November 2018-May 2019 (28 respondents). RESULTS: About 34.2% of respondents stated that tenofovir/emtricitabine (TDF/FTC) was licensed for use in their country in 2017, and 66.7% that it was licensed for use in 2018 (P = 0.02). PrEP was recommended in national guidelines in 39.5% of responses in 2017 and 40.7% in 2018 (P = 0.378). About 70.7% of respondents were aware of "informal" PrEP use in 2017, while 66.6% were aware of this in 2018 (P = 0.698). In 2018, there were 53 centres offering PreP (the highest numbers in Poland and Romania), whereas six countries had no centres offering PreP. The estimated number of HIV-negative people on PreP in the region was 4500 in 2018. Generic TDF/FTC costs (in Euros) ranged from €10 (Romania) to €256.92 (Slovakia), while brand TDF/FTC costs ranged from €60 (Albania) to €853 (Finland). CONCLUSIONS: Although the process of licensing TDF/FTC use for PrEP has improved, this is not yet reflected in the guidelines, nor has there been a reduction in the "informal" use of PrEP. PrEP remains a rarely used preventive method in CEE countries.

HIV care in times of the COVID-19 crisis - Where are we now in Central and Eastern Europe?

INTRODUCTION: The SARS-CoV-2 pandemic has hit the European region disproportionately. Many HIV clinics share staff and logistics with infectious disease facilities, which are now on the frontline in tackling COVID-19. Therefore, this study investigated the impact of the current pandemic situation on HIV care and continuity of antiretroviral treatment (ART) supplies in CEE countries. METHODS: The Euroguidelines in Central and Eastern Europe (ECEE) Network Group was established in February 2016 to review standards of care for HIV in the region. The group consists of professionals actively involved in HIV care. On March 19, 2020 we decided to review the status of HIV care sustainability in the face of the emerging SARS-CoV-2 pandemic in Europe. For this purpose, we constructed an online survey consisting of 23 questions. Respondents were recruited from ECEE members in 22 countries, based on their involvement in HIV care, and contacted via email. RESULTS: In total, 19 countries responded: Albania, Armenia, Belarus, Bosnia and Herzegovina, Bulgaria, Croatia, Czech Republic, Estonia, Georgia, Greece, Hungary, Lithuania, Macedonia, Poland, Republic of Moldova, Russia, Serbia, Turkey, and Ukraine. Most of the respondents were infectious disease physicians directly involved in HIV care (17/19). No country reported HIV clinic closures. HIV clinics were operating normally in only six countries (31.6%). In 11 countries (57.9%) physicians were sharing HIV and COVID-19 care duties. None of the countries expected shortage of ART in the following 2 weeks; however, five physicians expressed uncertainty about the following 2 months. At the time of providing responses, ten countries (52.6%) had HIV-positive persons under quarantine. CONCLUSIONS: A shortage of resources is evident, with an impact on HIV care inevitable. We need to prepare to operate with minimal medical resources, with the aim of securing constant supplies of ART. Non-governmental organizations should re-evaluate their earlier objectives and support efforts to ensure continuity of ART delivery.

Differentiation of Friend erythroleukemic cells in media lacking fetal calf serum.

The results presented in this paper provide evidence that the erythroid differentiation induced with n-butyric acid (BA) in F4N Friend cells depends mainly on the ability of different media to support an active cellular proliferation. No significant differences in the percentage of hemoglobin-positive cells were observed when a comparative induction was performed in media, supplemented with either calf or fetal calf serum; the most satisfactory results were obtained in Dulbecco's minimum essential medium (DMEM), lacking fetal calf serum but containing calf serum instead. The great sensitivity of cells to BA in the presence of calf serum allows us to suggest that there is not any component, specific for the fetal calf serum, critically required for the induction of erythroid differentiation in this clone.

The growth inhibition in Friend erythroleukemia cells induces cycling of preexisting nonhistone proteins.

The present investigation performed on Friend erythroleukemia cells provides evidence that the cellular reprogramming associated with transition of the cells from the growing to the resting state is accompanied by an intranuclear cycling of preexisting proteins migrating from the cytoplasm. The model system developed for this study affords an opportunity to follow these events in a conveniently short period. The flow microfluorometric analysis revealed that, with respect to DNA content, the quiescent nuclei exhibit distribution in G1, S and G2 phases, as do the nuclei of the control, exponentially growing cells. It was found further that the induction of quiescence arouses two waves of migration of preexisting cytoplasmic proteins toward the nucleus: an early one, during the transition to quiescence clearly showing density dependence, and a late one, in the established resting state. While the early-cycled proteins are undetectable in mass and once they have reached the nucleus have a short life time and, thus, could be considered as regulatory molecules, the late-cycled proteins accumulate within the nucleus and are associated most probably with structural reorganization of the resting nuclei; moreover, the late-cycled nonhistone proteins were found localized, at least partly, in the nuclear protein matrix structure. Control experiments confirmed that the early and late-cycled proteins have been synthesized in the cytoplasm in the proliferative state but are cycled intranuclearly only if the resting state has been induced. When finally the resting cells were stimulated to proliferate, the accumulated presynthesized and newly synthesized nonhistone proteins undergo a rapid degradation which suggests that the new cellular programme may require a complete elimination of the 'resting' nonhistone proteins.

Transformation suppressor activity of a Jun transcription factor lacking its activation domain.

The oncoprotein c-Jun is thought to be a mediator of ras transformation as both its synthesis and activity as a transcription factor are stimulated by ras expression. But c-Jun co-operates with ras in transformation assays, suggesting that they act along different pathways (reviewed in ref. 4). Here we show by means of a dominant-negative mutated transcription factor that c-Jun potentially in conjunction with other factors that interact with it is necessary for transformation by ras. The mutant Jun lacks an activation domain and blocks stimulation of transcription by several oncoproteins, including Ras, v-Src, polyoma middle T, c-Jun and c-Fos, as well as by the tumour promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). The inhibition is specific for motifs that bind Jun: activation of an NF-kappa B/Rel motif is not affected. This Jun mutant acts as an anti-oncogene in ras-transformed cells, generating non-transformed revertants that have acquired anchorage and density-dependent growth, as well as reduced tumorigenicity in vivo. Mutants of other transcription factors designed to inhibit transformation will enable us to study their role in signal transduction.

Evidence for the existence in the nuclei of Friend cells of a new type of ribonucleoprotein network.

We have isolated fragments of a novel nuclear structure exhibiting the morphological and biochemical characteristics of a ribonucleoprotein network. Under transmission electron microscopy it is visualized as irregularly interconnected branches assembled by tightly packed particles with sizes between 100A and 300A. RNA extracted from this structure shows a complex pattern: as well as ribosomal 28 S and 18 S RNA, small amounts of heterogeneous nuclear RNA and small nuclear RNA are also present. The protein composition of the network indicates a strong domination of ribosomal polypeptides. This fact, considered together with the sedimentation characteristics of the prevailing type of particles isolated directly from the network, supports the conclusion that ribosomal particles are the representative particulate elements. Further electrophoretic analysis of the protein has pointed out that it also contains a significant number of acidic polypeptides. Control experiments have suggested that the site of origin of the network fragments studied is not nucleolar: the fragments are released during extraction, most probably from the nuclear periphery. It is not yet clear whether this structure is localized only at the nuclear periphery or represents an extensive structure, occupying the entire volume of the nucleus. It is speculated that the network is involved in the extranucleolar transport and maturation of ribosomes.

Intranuclear distribution of the non-histone proteins: evidence for their compartmentalization.

1. Qualitative and quantitative distribution of the non-histone proteins in nuclear matrix, chromatin, a new type of RNP-network and nucleosol of Friend cells have been investigated. 2. The specific territorial distribution and metabolism of these proteins found support for the idea of their exact compartmentalization. 3. Since the majority of the non-histone proteins belong to the protein moiety of nuclear RNP-structures their specific territorial distribution probably express a primary compartmentalization of the nuclear ribonucleoproteins.

Histones of terminally differentiated cells undergo continuous turnover.

In contrast to the widely accepted idea of the nearly absolute metabolic stability of histones, our experiments support the view that the histones of nonproliferating, terminally differentiated cells undergo continuous replacement. This conclusion is based on the incorporation of labeled amino acids into the histones of mouse kidney and liver cells after their intraperitoneal introduction. We have found that the intranuclear uptake of the histones made in the absence of replicative synthesis and their integration into chromatin proceed with striking delay. The metabolic rates of individual histones measured by calculating their half-lives suggest that each histone turns over at a specific rate. With regard to the basic chromatin structure, the nucleosome, such unequal turnover should mean that the histone core does not participate in this process as a single unit but rather as a protein mosaic in which each partner follows its own rate of removal. Additional experiments suggested that intact nucleosomes take part in the replacement, but the relative proportion of the nucleosomes involved should be limited. The nonnucleosomal H1A and H1 degree histones have been found to undergo faster replacement than the core histones. Moreover, in comparison to each other, these two histone subfractions are also replaced at a different rate. The results of autoradiography of isolated kidney and liver nuclei after continuous labeling with [3H]-thymidine suggest that the histone replacement is not associated with the repair of DNA.

Increased proteolysis in chromatin of terminally differentiated and quiescent cells.

Endogenous proteolysis in chromatin of terminally differentiated, quiescent, and actively proliferating cells was studied by measuring the released acid-soluble radioactivity of [3H]tryptophan-prelabelled nuclear proteins, and by following the specific quantitative and qualitative changes in electrophoregrams of chromosomal proteins. The experiments suggest that the chromatin of differentiated mouse kidney and liver cells, as well as chromatin from Friend cells induced to commit terminal differentiation, exhibit increased proteolysis in comparison with that of chromatin isolated from actively proliferating cells. Enhanced proteolysis was found also for the slowly renewing and quiescent cells from adult mice. The control experiments designated to discriminate between the two possible alternatives explaining the difference--increased activity of the proteolytic enzymes associated with chromatin, or increased susceptibility of the chromosomal proteins to proteases--supported the latter alternative.