Nociceptor subtypes are born continuously over DRG development peaking at E10.5 - E11.5.

Sensory neurogenesis in the dorsal root ganglion (DRG) occurs in two waves of differentiation with larger, myelinated proprioceptive and low-threshold mechanoreceptor (LTMR) neurons differentiating before smaller, unmyelinated (C) nociceptive neurons. This temporal difference was established from early birthdating studies based on DRG soma cell size. However, distinctions in birthdates between molecular subtypes of sensory neurons, particularly nociceptors, is unknown. Here, we assess the birthdate of lumbar DRG neurons in mice using a thymidine analog, EdU, to label developing neurons exiting mitosis combined with co-labeling of known sensory neuron markers. We find that different nociceptor subtypes are born on similar timescales, with continuous births between E9.5 to E13.5, and peak births from E10.5 to E11.5. Notably, we find that thinly myelinated Aδ-fiber nociceptors and peptidergic C-fibers are born more broadly between E10.5 and E11.5 than previously thought and that non-peptidergic C-fibers and C-LTMRs are born with a peak birth date of E11.5. Moreover, we find that the percentages of nociceptor subtypes born at a particular timepoint are the same for any given nociceptor cell type marker, indicating that intrinsic or extrinsic influences on cell type diversity are occurring similarly across developmental time. Overall, the patterns of birth still fit within the classical ″two wave″ description, as touch and proprioceptive fibers are born primarily at E10.5, but suggest that nociceptors have a slightly broader wave of birthdates with different nociceptor subtypes continually differentiating throughout sensory neurogenesis irrespective of myelination.

Temporal Trends in HIV-1 Mutations Used for the Surveillance of Transmitted Drug Resistance

In 2009, a list of nonpolymorphic HIV-1 drug resistance mutations (DRMs), called surveillance DRMs (SDRMs), was created to monitor transmitted drug resistance (TDR). Since 2009, TDR increased and antiretroviral therapy (ART) practices changed. We examined the changing prevalence of SDRMs and identified candidate SDRMs defined as nonpolymorphic DRMs present on ≥ 1 expert DRM list and in ≥0.1% of ART-experienced persons. Candidate DRMs were further characterized according to their association with antiretrovirals and changing prevalence. Among NRTI-SDRMs, tenofovir-associated mutations increased in prevalence while thymidine analog mutations decreased in prevalence. Among candidate NRTI-SDRMs, there were six tenofovir-associated mutations including three which increased in prevalence (K65N, T69deletion, K70G/N/Q/T). Among candidate NNRTI-SDRMs, six that increased in prevalence were associated with rilpivirine (E138K/Q, V179L, H221Y) or doravirine (F227C/L) resistance. With the notable exceptions of I47A and I50L, most PI-SDRMs decreased in prevalence. Three candidate PI-SDRMs were accessory darunavir-resistance mutations (L10F, T74P, L89V). Adding the candidate SDRMs listed above was estimated to increase NRTI, NNRTI, and PI TDR prevalence by 0.1%, 0.3%, and 0.3%, respectively. We describe trends in the prevalence of nonpolymorphic HIV-1 DRMs in ART-experienced persons. These data should be considered in decisions regarding SDRM list updates and TDR monitoring.

Enhanced duplex- and triplex-forming ability and enzymatic resistance of oligodeoxynucleotides modified by a tricyclic thymine derivative

Oligodeoxynucleotides modified with a tricyclic thymidine analog (OBN) were synthesized, and their duplex- and triplex-forming ability, fluorescence properties and enzymatic stability were studied.

Toll-Like Receptor 2 Attenuates Traumatic Brain Injury-Induced Neural Stem Cell Proliferation in Dentate Gyrus of Rats

It was not clear how and whether neural stem cells (NSCs) responded to toll-like receptor 2 (TLR2) in the inflammatory environment after traumatic brain injury (TBI). The current study investigated the correlation of TLR2 and NSC proliferation in the dentate gyrus (DG) using the TBI model of rats. Immunofluorescence (IF) was used to observe the expression of BrdU, nestin, and TLR2 in the DG in morphology. Proliferating cells in the DG were labelled by thymidine analog 5-bromo-2-deoxyuridine (BrdU). Three-labelled BrdU, nestin, and DAPI was used for the identification of newly generated NSCs. Western blotting and real-time polymerase chain reaction (PCR) were used to observe the expression of TLR2 from the level of protein and mRNA. We observed that BrdU+/nestin+/DAPI+ cells accounted for 84.30%±6.54% among BrdU+ cells; BrdU+ and nestin+ cells in the DG were also TLR2+ cells. BrdU+ cells and the expression of TLR2 (both protein and mRNA levels) both elevated immediately at 6 hours (h), 24 h, 3 days (d), and 7 d posttrauma and peaked in 3 d. Results indicated that TLR2 was expressed on proliferating cells in the DG (NSCs possibly) and there was a potential correlation between increased TLR2 and proliferated NSCs after TBI. Taken together, these findings suggested that TLR2 was involved in endogenous neurogenesis in the DG after TBI.

Antiviral Activity of Tenofovir Alafenamide against HIV-1 with Thymidine Analog-Associated Mutations and M184V

ABSTRACT Tenofovir alafenamide (TAF) and tenofovir disoproxil fumarate (TDF) are prodrugs of the HIV-1 nucleotide reverse transcriptase inhibitor tenofovir (TFV). In vivo, TAF achieves >4-fold-higher intracellular levels of TFV diphosphate (TFV-DP) compared to TDF. Since thymidine analog-associated mutations (TAMs) in HIV-1 confer reduced TFV susceptibility, patients with TAM-containing HIV-1 may benefit from higher TFV-DP levels delivered by TAF. Moreover, the presence of the M184V mutation increases TFV susceptibility during TDF- or TAF-based therapy. The susceptibilities to antiviral drugs of site-directed mutants (SDMs) and patient-derived mutants containing combinations of TAMs (M41L, D67N, K70R, L210W, T215Y, and K219Q) with or without the M184V mutation (TAMs±M184V) were evaluated using either 5-day multicycle (MC; n = 110) or 2-day single-cycle (SC; n = 96) HIV assays. The presence of M184V in TAM-containing HIV-1 SDMs (n = 48) significantly increased TAF sensitivity compared to SDMs without M184V (n = 48). The comparison of TAF and TDF resistance profiles was further assessed in viral breakthrough (VB) experiments mimicking clinically relevant drug concentrations. A total of 68 mutants were assayed at physiological concentration in VB experiments, with 15/68 mutants breaking through with TDF (TFV, the in vitro equivalent of TDF, was used in these experiments), and only 3 of 68 mutants breaking through under TAF treatment. Overall, in the VB assay mimicking the 4-fold-higher intracellular levels of TFV-DP observed clinically with TAF versus TDF, TAF inhibited viral breakthrough of most TAM-containing HIV-1, whereas TDF did not. These results indicate that TAF has a higher resistance threshold than TDF and suggest that higher resistance cutoffs should be applied for TAF compared to TDF in genotypic and phenotypic resistance algorithms.