Distinct immunopathological mechanisms of EBV-positive and EBV-negative posttransplant lymphoproliferative disorders

EBV-positive and EBV-negative posttransplant lymphoproliferative disorders (PTLDs) arise in different immunovirological contexts and might have distinct pathophysiologies. To examine this hypothesis, we conducted a multicentric prospective study with 56 EBV-positive and 39 EBV-negative PTLD patients of the K-VIROGREF cohort, recruited at PTLD diagnosis and before treatment (2013–2019), and compared them to PTLD-free Transplant Controls (TC, n = 21). We measured absolute lymphocyte counts (n = 108), analyzed NK- and T cell phenotypes (n = 49 and 94), and performed EBV-specific functional assays (n = 16 and 42) by multiparameter flow cytometry and ELISpot-IFNγ assays (n = 50). EBV-negative PTLD patients, NK cells overexpressed Tim-3; the 2-year progression-free survival (PFS) was poorer in patients with a CD4 lymphopenia (CD4⁺<300 cells/mm³, p <  .001). EBV-positive PTLD patients presented a profound NK-cell lymphopenia (median = 60 cells/mm³) and a high proportion of NK cells expressing PD-1 (vs. TC, p = .029) and apoptosis markers (vs. TC, p < .001). EBV-specific T cells of EBV-positive PTLD patients circulated in low proportions, showed immune exhaustion (p = .013 vs. TC) and poorly recognized the N-terminal portion of EBNA-3A viral protein. Altogether, this broad comparison of EBV-positive and EBV-negative PTLDs highlight distinct patterns of immunopathological mechanisms between these two diseases and provide new clues for immunotherapeutic strategies and PTLD prognosis.     

A randomized controlled trial evaluating combination detection of HIV in Malawian sexually transmitted infections clinics

IntroductionHIV diagnosis is the necessary first step towards HIV care initiation, yet many persons living with HIV (PLWH) remain undiagnosed. Employing multiple HIV testing strategies in tandem could increase HIV detection and promote linkage to care. We aimed to assess an intervention to improve HIV detection within socio‐sexual networks of PLWH in two sexually transmitted infections (STI) clinics in Lilongwe, Malawi.MethodsWe conducted a randomized controlled trial to evaluate an intervention combining acute HIV infection (AHI) screening, contract partner notification and social contact referral versus the Malawian standard of care: serial rapid serological HIV tests and passive partner referral. Enrolment occurred between 2015 and 2019. HIV‐seropositive persons (two positive rapid tests) were randomized to the trial arms and HIV‐seronegative (one negative rapid test) and ‐serodiscordant (one positive test followed by a negative confirmatory test) persons were screened for AHI with HIV RNA testing. Those found to have AHI were offered enrolment into the intervention arm. Our primary outcome of interest was the number of new HIV diagnoses made per index participant within participants’ sexual and social networks. We also calculated total persons, sexual partners and PLWH (including those previously diagnosed) referred per index participant.ResultsA total of 1230 HIV‐seropositive persons were randomized to the control arm, and 561 to the intervention arm. Another 12,713 HIV‐seronegative or ‐serodiscordant persons underwent AHI screening, resulting in 136 AHI cases, of whom 94 enrolled into the intervention arm. The intervention increased the number of new HIV diagnoses made per index participant versus the control (ratio: 1.9; 95% confidence interval (CI): 1.2 to 3.1). The intervention also increased the numbers of persons (ratio: 2.5; 95% CI: 2.0 to 3.2), sexual partners (ratio: 1.7; 95% CI: 1.4 to 2.0) and PLWH (ratio: 2.3; 95% CI: 1.7 to 3.2) referred per index participant.ConclusionsCombining three distinct HIV testing and referral strategies increased the detection of previously undiagnosed HIV infections within the socio‐sexual networks of PLWH seeking STI care. Combination HIV detection strategies that leverage AHI screening and socio‐sexual contact networks offer a novel and efficacious approach to increasing HIV status awareness.     

A simple scoring system to estimate perioperative mortality following liver resection for primary liver malignancy—the Hepatectomy Risk Score (HeRS)

BackgroundSelection of the optimal treatment modality for primary liver cancers remains complex, balancing patient condition, liver function, and extent of disease. In individuals with preserved liver function, liver resection remains the primary approach for treatment with curative intent but may be associated with significant mortality. The purpose of this study was to establish a simple scoring system based on Model for End-stage Liver Disease (MELD) and extent of resection to guide risk assessment for liver resections.MethodsThe 2005–2015 NSQIP database was queried for patients undergoing liver resection for primary liver malignancy. We first developed a model that incorporated the extent of resection (1 point for major hepatectomy) and a MELD-Na score category of low (MELD-Na =6, 1 point), medium (MELD-Na =7–10, 2 points) or high (MELD-Na >10, 3 points) with a score range of 1–4, called the Hepatic Resection Risk Score (HeRS). We tested the predictive value of this model on the dataset using logistic regression. We next developed an optimal multivariable model using backwards sequential selection of variables under logistic regression. We performed K-fold cross validation on both models. Receiver operating characteristics were plotted and the optimal sensitivity and specificity for each model were calculated to obtain positive and negative predictive values.ResultsA total of 4,510 patients were included. HeRS was associated with increased odds of 30-day mortality [HeRS =2: OR =3.23 (1.16–8.99), P=0.025; HeRS =3: OR =6.54 (2.39–17.90), P<0.001; HeRS =4: OR =13.69 (4.90–38.22), P<0.001]. The AUC for this model was 0.66. The AUC for the optimal multivariable model was higher at 0.76. Under K-fold cross validation, the positive predictive value (PPV) and negative predictive value (NPV) of these two models were similar at PPV =6.4% and NPV =97.7% for the HeRS only model and PPV =8.4% and NPV =98.1% for the optimal multivariable model.ConclusionsThe HeRS offers a simple heuristic for estimating 30-day mortality after resection of primary liver malignancy. More complicated models offer better performance but at the expense of being more difficult to integrate into clinical practice.     

A pilot study of cadre training to promote responsible self-medication in Indonesia: Which is better specific or general modules?

In 2015, the Indonesian Government initiated ‘Smart Use of Medications Movement’ (‘GeMa CerMat’) which included cadre training to promote responsible self-medication. Evaluation of a pilot training conducted across Indonesia suggested the need to improve those training modules. This study aimed to assess cadre’ knowledge gained following training with newly developed general or specific training modules. Five types of modules were developed and used to train cadres at five Community Health Centres (CHCs) in Surabaya, Indonesia: 1) Sidosermo CHC (general-drugs module), 2) Tenggilis CHC (common cold drugs module), 3) Gunung Anyar CHC (analgesic drugs module), 4) Kalirungkut CHC (anti-diarrhoeal drugs module), and 5) Jagir CHC (indigestion drugs module). Cadres’ knowledge improvements were evaluated using pre-/post-test scores and the difference scores depending on the module being tested. Multifactorial ANOVA explored the effects of the type of module on difference scores. A total of 279 cadres across five CHCs were involved in the training, giving response rates from 65% to 93%. There was an increase in the post-test scores after the training with all modules. However, significant differences were reported only for the specific-drugs module groups (all p < .001). Furthermore, the general module group had the lowest difference score (1.12; 95% CI [−0.45, 2.92]) while the common cold module group had the highest gain (5.02; 95% CI [1.95, 5.17]). Multifactorial ANOVA revealed that there was a significant main effect of the type of modules on difference scores [F (4, 263) = 8.37, p <.001]. In conclusion, this preliminary study indicated that the development of modules for specific minor illnesses could be beneficial in facilitating effective community-based training to promote responsible self-medication in Indonesia. The priority for therapeutic areas chosen for the module should be based on the local needs. Further research is required to confirm the findings in broader community members.     

Protein binding of brequinar in the plasma of healthy donors and cancer patients and analysis of the relationship between protein binding and pharmacokinetics in cancer patients

The protein binding of weakly acidic and basic drugs has been shown to be altered in cancer patients. Brequinar is a weakly acidic, low-clearance, and highly protein-bound (>98% bound) antitumor agent. The pharmacokinetic parameters of brequinar are subject to large interpatient variability. This large interpatient variability may be related to brequinar's plasma protein-binding capacity (assuming no change in the intrinsic clearance of the unbound drug). The objectives of this study, therefore, were (a) to characterize brequinar's protein binding in the plasma of healthy donors and cancer patients and (b) to examine the relationships between brequinar's plasma protein binding and its pharmacokinetics in patients. Brequinar protein binding was determined in human serum albumin (HSA) solution, drug-free donor plasma, and brequinar-free, predose plasma samples obtained from a phase I cancer trial. Pharmacokinetic results from this study were used to examine relationships between plasma protein binding and drug disposition. In HSA solution and healthy donor plasma, brequinar's protein binding as determined using spiked samples was concentration-dependent. The unbound brequinar fraction increased by a factor of 3 (from 0.3% to 0.9% free) in 4% HSA solution and by a factor of 4 (from 0.4% to 1.6% free) in donor plasma as the brequinar concentrations increased from 0.1 to 2.3 mM in the HSA solution and from 0.076 to 1.5 mM in the donor plasma. Analysis of brequinar binding characteristics using the binding ratio and Rosenthal binding plots showed that albumin was the primary protein for brequinar binding in human plasma. The addition of various concentrations of ₁-acid glycoprotein to 4% HSA solution did not affect the protein binding of brequinar to HSA. The protein binding determined in the plasma of cancer patients was not quantitatively different, except for variability, from that observed in the plasma of healthy donors. Examination of relationships between the unbound brequinar fraction and pharmacokinetics suggested that plasma protein binding was not a major determinant of brequinar disposition in cancer patients.     

Ovulation triggering in clomiphene citrate-stimulated cycles: Human chorionic gonadotropin versus a gonadotropin releasing hormone agonist

Purpose To compare the use of human chorionic gonadotropin (hCG) to a gonadotropin releasing hormone (GnRH) agonist, nafarelin, in initiating ovulation and supporting the luteal phase after priming with clomiphene.Methods In 26 infertile women 50 mg clomiphene citrate produced a preovulatory-size follicle. Then, 11 women were randomized to receive two 400-g doses of nafarelin intranasally 16 h apart, and 15 women were injected intramuscularly with 5000 IU of hCG (luteal day 0 = LD0). Starting on LD6, 7 more 400-g doses of nafarelin were repeated on an every 16-h schedule or a single 2500 IU dose of hCG was given, respectively. Serum levels of follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E₂), progesterone (P), and hCG were measured. On LD13, endometrium was evaluated with ultrasonography and biopsy in 19 nonpregnant women.Results As judged by a threefold rise in serum LH, an LH surge was detected on LD1 in all 11 nafarelin patients, but in only 8 hCG patients (P = 0.01). LH and FSH levels were significantly higher on LD1, 7, and 8 and were significantly suppressed on LD13 in the nafarelin group. All patients had mid-luteal P levels greater than 10 ng/ml and luteal phases longer than 13 days. Significantly different luteal E₂ or P levels were noted only on LD13, with lower values in the nafarelin group. Pregnancies were achieved in 3 of 11 nafarelin cycles and 2 of 15 hCG cycles. Luteal phase defects were also similar: 4 of 8 nafarelin patients and 7 of 11 hCG patients.Conclusion Nafarelin or hCG in conjunction with clomiphene can result in viable pregnancies, but is associated with low pregnancy rates and a high incidence of luteal phase defects.     

Pax-1 in the development of the cervico-occipital transitional zone

The Pax-1 gene has been found to play an important role in the development of the vertebral column. The cervico-occipital transitional zone is a specialized region of the vertebral column, and malformations of this region have frequently been described in humans. The exact embryonic border between head and trunk is a matter of controversy. In order to determine a possible role of Pax-1 in the development of the cervico-occipital transitional zone we studied the expression of this gene in a series of quail embryos and murine fetuses with in situ hybridization and immunohistochemistry. Pax-1 is expressed in all somites of the embryo, including the first five occipital ones. During embryonic days 3–5 the gene is down-regulated in the caudal direction within the first five somites, whereas more caudally Pax-1 is strongly expressed in the cells of the perinotochordal tube. In 5-day-old quail embryos, the cartilaginous anlage of the basioccipital bone has developed and there is no more expression of Pax-1 in this region. The fusion of the dens axis with the body of the axis also coincides with switching off of the Pax-1 gene. More caudally, the gene is continuously expressed in the intervertebral discs of murine embryos and therefore seems to be important for the process of resegmentation. Quail embryos do not possess permanent intervertebral discs. Hyper- or hyposegmentation defects may be explained by an over- or under-expression of Pax-1 during development. We also reinvestigated the border between the head and trunk in chick embryos by performing homotopical grafting experiments of the 5^th somite between chick and quail embryos. Grafted quail cells formed mainly the caudal end of the basioccipital bone. They were also located in the cranial half of the ventral atlantic arch, and only a few cells were found in the tip of the dens axis.     

Mutational specificity of aflatoxin B1. Comparison of in vivo host-mediated assay with in vitro S9 metabolic activation

An intrasanguineous host-mediated assay was used to determinethe pattern of mutagenesis induced by the carcinogen aflatoxinBl in the lacl gene of Escherichia coli recovered from rat liver.To investigate the influence of different types of metabolicactivation, the mutation spectrum induced by AFB1 activatedin vitro by a commercially prepared S9 microsomal fraction fromAroclor 1254-treated rats was also obtained. A total of 281forward mutations affecting the N-terminal region of the laclgene were characterized by DNA sequencing analysis. AFB1 inducedsimilar type of mutations with similar site specificity whenactivated by the standard S9 fraction or by employing a rathost-mediated assay. These results indicate the ability of thein vitro S9 fraction to mimic the in vivo metabolism, suggestingthat the same active metabolite, presumably AFB1 8, 9-epoxide,is responsible for generating a similar pattern of DNA damage,as reflected in the similarity of mutational spectra. For bothactivation systems, most mutations (>90%) were base substitutionsthat occurred primarily at G: C pairs. Somewhat over one-halfof G: C targeted substitutions were GC>TA transversions,other mutations being evenly divided between G: C>AT transitionsand GC>CG trans-versions. The mutational specificity exhibitedby activated AFB1 can be explained by incorporation of differentbases opposite a single type of non-instructive lesion duringerror-prone DNA synthesis. To what extent the mutations aredue to the main adduct (AFB1-N7-Gua), its imidazole-ring-openedderivative or an apurinic site remains unknown.     

Gallium scintigraphy in Hansen's disease

Gallium 67 imaging was used in 12 patients with documented Hansen's disease undergoing treatment or not, in an attempt to determine the pattern of the disease. Diagnosis was confirmed by histopathology in all patients. The Mitsuda reaction was seen in all patients. Specific nuclear studies were performed when needed to evaluate particular organs better. Gallium 67 images show homogeneous, diffuse and moderate accumulation over the entire skin surface (except for the face) of untreated patients with multibacillary disease. The facial skin in these cases presented homogeneous, diffuse but very marked uptake of gallium. Internal organ involvement was variable. There was a very good correlation among clinical, scintigraphic, immunological and histopathological data. The pattern of the body skin (skin outlining) and facial skin (beard distribution) may be distinct for untreated patients with multibacillary leprosy.     

Seroreactivity to human papillomavirus (HPV) types 16, 18, or 31 and risk of subsequent HPV infection: results from a population-based study in Costa Rica.

Whether antibodies to human papillomavirus (HPV) capsids, elicited by natural infection, are protective is unknown. This question was addressed in a population-based cohort of 7046 women in Costa Rica by examining the association between baseline seroreactivity to HPV-16, HPV-18, or HPV-31 virus-like particles and the risk of subsequent HPV infection at a follow-up visit 5-7 years after enrollment. Seropositivity to HPV-16, HPV-18, or HPV-31 was not associated with a statistically significant decreased risk of infection with the homologous HPV type [relative risk (RR) and [95% confidence interval (CI)], 0.74 (0.45-1.2), 1.5 (0.83-2.7), and 0.94 (0.48-1.8), respectively]. Seropositivity to HPV-16 or HPV-31 was not associated with a decreased risk of infection with HPV-16 or its genetically related types [RR (95% CI), 0.82 (0.61-1.1) and 0.93 (0.68-1.2), respectively]. Seropositivity to HPV-18 was not associated with a decreased risk of infection with HPV-18 or its genetically related types (RR 1.3; 95% CI 1.0-1.8). Thus, we did not observe immunity, although a protective effect from natural infection cannot be excluded because of the limits of available assays and study designs.