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sparser
"The TRPV1 receptor and Interleukin-1β (IL-1β) are key upstream triggering factors in the Cav3.2/USP5 pain pathway, and macrophage-derived extracellular vesicles containing miR-23a-3p and SUMOylation c[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"

sparser
"New analgesics have been developed that target the USP5/Cav3.2 channel, including Suramin (an anti-trypanosomal and anti-filarial drug), Flavonoid, and Gossypetin, which inhibit the binding of USP5 an[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"

sparser
"Rodent compound II-1 inhibits the biochemical interaction between USP5 and Cav3.2 III-IV junctions in a dose-dependent manner and suppresses mechanical abnormal pain caused by sciatic nerve injury [ 9[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"

sparser
"Lamina II (substantia gelatinosa (SG)) neurons in the superficial spinal dorsal horn (SDH) are the second-order sensory neurons, which primarily receive and modulate nociceptive inputs from the myelinated Aδ and unmyelinated C fibers. xref Therefore, the increasing excitatory process of SG neuron is one of the most important mechanisms for the central sensitization of pathological pain. xref Recently, a Cav3.2-dependent link of SG neuronal excitability and cutaneous transient receptor potential vanilloid-1 (TRPV1) expressing nociceptors has been demonstrated. xref Conditioning stimulation of optogenetically targeted cutaneous TRPV1 expressing nociceptors could evoke an increase in C-fiber activity, which may, in turn, lead to an enhanced presynaptic excitatory neurotransmission in SG neurons. xref Moreover, either Ni 2+ (a blocker of Cav3 channels) or Tat-3.2-III-IV peptide (a blocker of USP5-Cav3.2 association) could decrease the frequency of excitatory postsynaptic currents in SG neurons, strongly suggesting a role of Cav3.2 in the sensitization of the pain pathway mentioned above. xref , xref Therefore, the potentiation of presynaptic Cav3.2 channels may contribute to the hyperexcitability of SG neurons, accompanying with the promotion of chronic pain."

sparser
"Icariside II (ICA-II), one of the flavonols studied, inhibited the biochemical interactions between USP5 and Cav3.2 and concomitantly and effectively blocked Cav3.2 channels."

sparser
"Icariside II, a Prenyl-Flavonol, Alleviates Inflammatory and Neuropathic Pain by Inhibiting T-Type Calcium Channels and USP5-Cav3.2 Interactions."

reach
"Current findings related to the small molecules suramin (IC 50 = ∼0.8 μM) and gossypetin (IC 50 = ∼20 μM) suggest that blocking the interaction between USP5 and Cav3.2 potentially opens an avenue for [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"

reach
"Disrupting the interaction between Cav3.2 and USP5 using the TAT-cUBP1-USP5 peptide has been shown to reduce the levels of the Cav3.2 calcium channel in vitro, which attenuates thermal hyperalgesia in diabetic neuropathy animals (137, 138)."

sparser
"A further study identified interleukin-1 β as the dominant driver and mediator of the elevated Cav3.2-USP5 interaction in the pain pathway [ 58 ]."

sparser
"These results indicate that blocking the USP5-Cav3.2 interaction is a potential strategy for treating pain."

sparser
"Screening for modulators of the USP5-Cav3.2 interaction."

sparser
"Current findings related to the small molecules suramin (IC 50 = ∼0.8 μM) and gossypetin (IC 50 = ∼20 μM) suggest that blocking the interaction between USP5 and Cav3.2 potentially opens an avenue for [MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"

sparser
"Here, we describe the development of an ELISA-based assay to screen a library of pharmacologically active compounds (including clinically used drugs) for molecules capable of disrupting the USP5-Cav3.2 interaction."

sparser
"While our Tat peptide approach delivered proof of concept for targeting the Cav3.2-USP5 interaction as a strategy for targeting various pain conditions, small organic mimetics of these peptides are a preferred strategy for therapeutics."

sparser
"We therefore developed an ELISA-based assay for identifying small organic disruptors of the Cav3.2-USP5 interaction."

sparser
"Suramin and gossypetin inhibited USP5 binding to Cav3.2 channels by 50-60% at 5 μM (Figure  xref A-D)."

sparser
"Altogether, these data reveal suramin and gossypetin as potential small organic disruptors of USP5 interactions with Cav3.2."

sparser
"No significant effects on current amplitude or biophysical properties of the channel were observed (7±2% inhibition (n=3) and 3±1% inhibition (n=3) of channel activity for suramin and gossypetin, respectively, which is indistinguishable from rundown), consistent with an action on the USP5-Cav3.2 channel interaction rather than direct effects on Cav3.2 channel function."

reach
"The acute sensitization initiated by optical stimulation (10 Hz) of TRPV1-ChR2 neurons, is sufficient to increase USP5 expression, which results in increased Cav3.2 T-type activity and increased mechanical hypersensitivity that is dependent on the interaction of USP5 and Cav3.2 [104]."

sparser
"Next, we determined if suramin altered endogenous USP5-Cav3.2 protein interactions in the dorsal horns from diabetic and non-diabetic mice."

sparser
"Suramin inhibited USP5 binding to Cav3.2 channels obtained from ob/ob mouse dorsal horns while total Cav3.2 protein levels remained similar (Figure  xref B, C)."

sparser
"The biflavenoid gossypetin (318 Da) and the polysulphonated naphtylurea suramin (~1.3 kDa) are vastly different in size, and yet both compounds effectively disrupted USP5 binding to the Cav3.2 III-IV linker region."

reach
"Here, we report the effects of SUMOylation on USP5 interactions with Cav3.2 calcium channels."

reach
"Overall, our observations extend our previous findings showing that peripheral nerve injury upregulates USP5 levels and leads to an enhanced interaction between USP5 and Cav3.2 [6]."

reach
"Decreased SUMOylation of USP5 after nerve injury would aid this process such that USP5 interactions with Cav3.2 are enhanced when USP5 SUMOylation is decreased."

sparser
"The latter is reminiscent of the hydroxylated chromene core of gossypetin, thus suggesting the possibility that the disruption of USP5-Cav3.2 interactions may involve this bicyclic aromatic structure."

sparser
"Overall, our results indicate that disrupting the interaction between the deubiquitinase USP5 and Cav3.2 calcium channels via a small organic molecule is a promising new strategy for treating a spectrum of chronic pain states."

sparser
"Altogether these data indicate that non-invasive trans-cutaneous activation of nociceptors triggers an upregulation of Cav3.2-USP5 complexes in both DRG and spinal dorsal horn."

sparser
"We next disrupted USP5 interactions with Cav3.2 by pretreating spinal cord slices with Tat-3.2-III-IV peptide."

reach
"We have shown previously that the first half of the cUBP domain of USP5 is the key structural determinant of USP5 binding to Cav3.2 [8]."

sparser
"Figure 3C Optogenetically Induced Heightening of Pain Responses Rely on USP5-Cav3.2 Interactions (corrected) Figure 3C Optogenetically Induced Heightening of Pain Responses Rely on USP5-Cav3.2 Interac[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"

reach
"We have previously reported that pain hypersensitivity in diabetic ob/ob mice can be reversed by blocking the interactions between USP5 and Cav3.2 by small organic molecule mimetics."

sparser
"Molecular docking analysis reveals two potential binding pockets at the USP5-Cav3.2 interface that are distinct from the binding site of the deubiquitinase inhibitor WP1130 (a."

reach
"Impairment of USP5 and Cav3.2 interaction attenuated inflammatory and neuropathic pain (109)."

reach
"IL-1β was an essential mediator to regulate the interaction between Cav3.2 and USP5 in the pain process (111)."

reach
"Impairing the binding between USP5 and Cav3.2 protected mechanical hypersensitivity in female mice with peripheral inflammation (112)."

sparser
"Uncoupling the Cav3.2-USP5 interaction produces analgesia in vivo , and these advances pave the way for other conceptually similar approaches that focus on a divergent molecular target [ xref – xref ]."

sparser
"Moreover, IL-1β might be an upstream trigger for the upregulation of Cav3.2-USP5 interactions in the pain pathway by increased firing activity in afferent fibers [14] ."

reach
"It has also been reported to have anti -hyperalgesia activity in treating neuropathic pain by disrupting protein-protein interaction between USP5, a deubiquitinating enzyme, and Cav3.2, a T-type calci[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"

sparser
"The deubiquitinase USP5 interacts with Cav3.2 and increases its stability and activity by removal of ubiquitin modification from Cav3.2."

sparser
"A novel bioactive compound (II-1) selectively binds to the interface of USP5-Cav3.2 and disrupts the interaction without affecting the function of USP5 on other targets [158] ."

sparser
"Suramin and the flavonoid gossypetin, the small molecules as USP5-Cav3.2 disruptors, produced dose-dependent and long-lasting mechanical anti-hyperalgesia in mouse models of neuropathic pain [82] ."

reach
"We showed that Cav3.2 channels associate with USP5, a deubiquitinating enzyme [16,17] that is upregulated in dorsal root ganglion (DRG) and dorsal horn tissue in response to either nerve injury or peripheral inflammation."

sparser
"The acute sensitization initiated by optical stimulation (10 Hz) of TRPV1-ChR2 neurons, is sufficient to increase USP5 expression, which results in increased Cav3.2 T-type activity and increased mechanical hypersensitivity that is dependent on the interaction of USP5 and Cav3.2 [ xref ]."

reach
"To detect USP5 bound to this immobilized Cav3.2 III-IV linker peptide we used a rabbit polyclonal anti-USP5 antibody, followed by the addition of an HRP-conjugated secondary antibody."

reach
"Suramin and gossypetin inhibited USP5 binding to Cav3.2 channels by 50-60% at 5 μM (Figure 3A-D)."

reach
"Altogether, these data reveal suramin and gossypetin as potential small organic disruptors of USP5 interactions with Cav3.2."

reach
"Suramin inhibited USP5 binding to Cav3.2 channels obtained from ob/ob mouse dorsal horns while total Cav3.2 protein levels remained similar (Figure 8B, C)."

reach
"Icariside II (ICA-II), one of the flavonols studied, inhibited the biochemical interactions between USP5 and Cav3.2 and concomitantly and effectively blocked Cav3.2 channels."

reach
"USP5 levels as well as the association between USP5 and Cav3.2 were upregulated in DRG on the side ipsilateral to stimulation ( Figures 2 A and 2B )."

reach
"Over a 24-hr period, however, there was a time-dependent waning of the thermal and mechanical hypersensitivity in opto-stimulated TRPV1-ChR2-EYFP animals ( Figures S2 A and S2B, respectively), which w[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"

reach
"Taken together, we attribute the increase in excitatory neurotransmission to a primarily presynaptic phenomenon.We next disrupted USP5 interactions with Cav3.2 by pretreating spinal cord slices with T[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"

sparser
"To determine which part of USP5 interacts with Cav3.2 channels, we designed short peptides (35–45 mers) corresponding to each of the major USP5 domains xref , with each peptide containing at least one α-helix."

sparser
"We have previously reported that pain hypersensitivity in diabetic ob/ob mice can be reversed by blocking the interactions between USP5 and Cav3.2 by small organic molecule mimetics. xref To determine whether the TAT-cUBP1-USP5 peptide was similarly effective, we assessed thermal withdrawal threshold in ob/ob mice before and after delivery of the TAT-cUBP1-USP5 peptide."

sparser
"Given the relevance of the pro-inflammatory cytokine interleukin-1 beta in many forms of pathological pain, we hypothesized that interleukin-1 beta may be a critical cofactor required to drive upregulation of interactions between USP5 and Cav3.2 channels."

sparser
"Consistent with our observations, this region has been previously described as a potential site for substrate targeting and specificity. xref In contrast, this region does not appear to be important for substrate modification by the catalytic site because deletion of the cUBP (163–291) domain did not affect the hydrolysis rate of ubiquitin-AMC. xref Together with our previous identification of a short (∼20 amino acid) stretch of residues in the domain III-IV linker of the Cav3.2 channel, xref we now have two complementary tools that allow us to disrupt USP5-Cav3.2 interactions both in vitro and in vivo."

sparser
"In summary, we have extended our previous work on USP5-Cav3.2 channel interactions to identify a key structural USP5 domain that is responsible for the actions of USP5 on this channel isoform."

sparser
"In contrast, co-immunoprecipitation experiments revealed that intrathecal administration of interleukin-1 beta in wild-type mice led to an increase in the interaction between USP5 and Cav3.2 in the spinal dorsal horn."

sparser
"Altogether, our findings identify interleukin-1 beta as an upstream trigger for the upregulation of interactions between USP5 and Cav3.2 channels in the pain pathway, presumably by triggering increased firing activity in afferent fibers."

sparser
"Since this process is relevant to both inflammatory and neuropathic pain states, xref , xref we initially proposed that neural activity may be a common trigger. xref This idea was supported by observations that activity stemming from optogenetically targeted cutaneous nociceptors alone is sufficient to upregulate USP5 expression, and therefore to increase interactions between USP5 and Cav3.2. xref However, this activity-dependent phenomenon is not self-sustaining, instead promoting only a transient pain state that cannot account for the upregulation observed after injury."

sparser
"t. injection of IL-1β (0.1 pg), co-immunoprecipation (co-IP) experiments demonstrated that IL-1β increases the interaction between USP5 and Cav3.2 in the spinal dorsal horn, relative to vehicle control (PBS) mice ( xref and ( xref ))."

reach
"It was recently demonstrated that this nociceptive effect is due to an increase in channel activity, which is primarily mediated by an upregulation of USP5, a deubiquitinating enzyme, which associates with CaV3.2 and inhibits its degradation [41]."

reach
"Using an intrathecal injection of a permeant peptide which impairs the interaction between USP5 and CaV3.2, several studies showed that it was possible to relieve rapidly (in a few minutes) pain sensation related to mechanical or thermal hypersensitivity.As a conclusion, many approaches using permeant peptides but also nonpeptidic synthetic molecules [42,43] have managed to alter the interaction between voltage-dependent calcium channel partners in order to successfully reverse various symptoms associated with different pathologies."

sparser
"While the above findings support the hypothesis that IL-1β drives upregulation of T-type channels via USP5 to induce pain, these effects were studied under acute conditions (<1 h) that yield transient pain (∼15 min) and may be secondary to interactions between IL-1β and glial cells, which are known to express IL-1RI. xref , xref To study neuroinflammatory interaction more directly, while reflecting sustained release after physical injury, we used DRG neuron cultures to evaluate the level of USP5 bound to Cav3.2 in response to overnight (∼24 h) exposure to IL-1β."

reach
"In contrast, co-immunoprecipitation experiments revealed that intrathecal administration of interleukin-1 beta in wild-type mice led to an increase in the interaction between USP5 and Cav3.2 in the spinal dorsal horn."

reach
"This idea was supported by observations that activity stemming from optogenetically targeted cutaneous nociceptors alone is sufficient to upregulate USP5 expression, and therefore to increase interactions between USP5 and Cav3.2."

sparser
"Under these conditions and relative to vehicle control (0.1% BSA in PBS), IL-1β produced a clear increase in the level of interaction between USP5 and Cav3.2 channels in co-IP experiments ( xref and ( xref )), which is mediated by IL-1RI, as this effect was almost completely attenuated in the presence of IL-1Ra (100 ng/ml)."

reach
"Moreover, disruption of the interaction between USP5 and Cav3.2 with TAT peptides suppressed acute nocifensive responses produced by interleukin-1 beta, which was similar to that achieved by elimination of T-type channel activity with the channel blockers, mibefradil, or TTA-A2."

reach
"injection of IL-1β (0.1 pg), co-immunoprecipation (co-IP) experiments demonstrated that IL-1β increases the interaction between USP5 and Cav3.2 in the spinal dorsal horn, relative to vehicle control (PBS) mice (Figure 1(b) and (c))."

reach
"To study neuroinflammatory interaction more directly, while reflecting sustained release after physical injury, we used DRG neuron cultures to evaluate the level of USP5 bound to Cav3.2 in response to overnight (∼24 h) exposure to IL-1β."

reach
"In support of such a mechanism, we discovered that, by sustaining elevated levels of IL-1β in DRG cultures, interactions between USP5 and Cav3.2 can be maintained."

reach
"It is interesting that in a preliminary in vitro screen of potential mediators, which included brain-derived neurotrophic factor, nerve growth factor, and tumor necrosis factor-alpha, none were as effective as IL-1β at inducing interactions between USP5 and Cav3.2 (data not shown)."

sparser
"Therefore, through disruption, the interaction between USP5 and Cav3.2 channel inhibits the expression of Cav3.2; hence, inhibiting T-type calcium current may alleviate NP ( xref )."

sparser
"When the SUMOylation of USP5 is decreased, there is an increase in the interactions between USP5 and Cav3.2 calcium channels. xref The KRas indirectly regulates USP5 activity by upregulating cellular ROS levels, leading to the formation of USP5 homodimers, enhanced USP5 enzymatic activity, and the accumulation of USP5 protein. xref Long non-coding RNAs (LncRNAs) interact with specific proteins to regulate downstream effectors."

sparser
"In support of such a mechanism, we discovered that, by sustaining elevated levels of IL-1β in DRG cultures, interactions between USP5 and Cav3.2 can be maintained."

sparser
"Some factors also affect the interaction between USP5 and Cav3.2 channels, such as IL-1 recognition ( xref ), USP5 SUMOylation disorder ( xref ), and TRPV1 nociceptors ( xref )."

sparser
"It is interesting that in a preliminary in vitro screen of potential mediators, which included brain-derived neurotrophic factor, nerve growth factor, and tumor necrosis factor-alpha, none were as effective as IL-1β at inducing interactions between USP5 and Cav3.2 (data not shown)."

sparser
"Our study focuses on disrupting the Cav3.2-USP5 interaction as a strategy for chronic pain management."

sparser
"Through structure-activity relationship studies of a tetrahydroquinoline (THQ) scaffold, we identified a family of lead molecules that demonstrated potent inhibition of the Cav3.2-USP5 interaction."

sparser
"Disrupting the interaction between Cav3.2 and USP5 using the TAT-cUBP1-USP5 peptide has been shown to reduce the levels of the Cav3.2 calcium channel in vitro , which attenuates thermal hyperalgesia in diabetic neuropathy animals ( xref , xref )."

sparser
"In addition, TRPV1, IL-1β, and HMGB1 activate the deubiquitinating enzyme USP5, which interacts with Cav3.2 and enhances whole-cell currents [ xref ]."

reach
"New analgesics have been developed that target the USP5/Cav3.2 channel, including Suramin (an anti-trypanosomal and anti-filarial drug), Flavonoid, and Gossypetin, which inhibit the binding of USP5 an[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]"

sparser
"Here we describe the regulation of the Cav3.2-USP5 interaction by SUMOylation."

sparser
"Here, we report the effects of SUMOylation on USP5 interactions with Cav3.2 calcium channels."

sparser
"A Synthetically Accessible Small-Molecule Inhibitor of USP5-Cav3.2 Calcium Channel Interactions with Analgesic Properties."

sparser
"We then sought to determine whether SUMOylation of USP5 affects the interaction between USP5 and Cav3.2 channels."

sparser
"This experiment suggests that SUMOylation state of USP5 can regulate USP5 interactions with Cav3.2 calcium channels."

sparser
"Overall, our observations extend our previous findings showing that peripheral nerve injury upregulates USP5 levels and leads to an enhanced interaction between USP5 and Cav3.2 [ xref ]."

sparser
"Decreased SUMOylation of USP5 after nerve injury would aid this process such that USP5 interactions with Cav3.2 are enhanced when USP5 SUMOylation is decreased."

sparser
"Hence, preventing deSUMOylation of USP5 could provide a strategy for enhancing USP5 interactions with Cav3.2, which would in turn be predicted to lead to analgesia."

sparser
"Notably, quercetin was found to significantly disrupt the interaction between USP5 (deubiquitinase) and Cav3.2."

sparser
"IL-1β was an essential mediator to regulate the interaction between Cav3.2 and USP5 in the pain process ( xref )."