Amphiregulin Switches Progenitor Cell Fate for Lineage Commitment During Gastric Mucosal Regeneration.

BACKGROUND & AIMS: Isthmic progenitors, tissue-specific stem cells in the stomach corpus, maintain mucosal homeostasis by balancing between proliferation and differentiation to gastric epithelial lineages. The progenitor cells rapidly adopt an active state in response to mucosal injury. However, it remains unclear how the isthmic progenitor cell niche is controlled during the regeneration of damaged epithelium. METHODS: We recapitulated tissue recovery process after acute mucosal injury in the mouse stomach. Bromodeoxyuridine incorporation was used to trace newly generated cells during the injury and recovery phases. To define the epithelial lineage commitment process during recovery, we performed single-cell RNA-sequencing on epithelial cells from the mouse stomachs. We validated the effects of amphiregulin (AREG) on mucosal recovery, using recombinant AREG treatment or AREG-deficient mice. RESULTS: We determined that an epidermal growth factor receptor ligand, AREG, can control progenitor cell lineage commitment. Based on the identification of lineage-committed subpopulations in the corpus epithelium through single-cell RNA-sequencing and bromodeoxyuridine incorporation, we showed that isthmic progenitors mainly transition into short-lived surface cell lineages but are less frequently committed to long-lived parietal cell lineages in homeostasis. However, mucosal regeneration after damage directs the lineage commitment of isthmic progenitors towards parietal cell lineages. During recovery, AREG treatment promoted repopulation with parietal cells, while suppressing surface cell commitment of progenitors. In contrast, transforming growth factor-α did not alter parietal cell regeneration, but did induce expansion of surface cell populations. AREG deficiency impairs parietal cell regeneration but increases surface cell commitment. CONCLUSIONS: These data demonstrate that different epidermal growth factor receptor ligands can distinctly regulate isthmic progenitor-driven mucosal regeneration and lineage commitment.

Rab11FIP1-deficient mice develop spontaneous inflammation and show increased susceptibility to colon damage.

The small GTPase, Rab11a, regulates vesicle trafficking and cell polarity in epithelial cells through interaction with Rab11 family-interacting proteins (Rab11-FIPs). We hypothesized that deficiency of Rab11-FIP1 would affect mucosal integrity in the intestine. Global Rab11FIP1 knockout (KO) mice were generated by deletion of the second exon. Pathology of intestinal tissues was analyzed by immunostaining of colonic sections and RNA-sequencing of isolated colonic epithelial cells. A low concentration of dextran sodium sulfate (DSS, 2%) was added to drinking water for 5 days, and injury score was compared between Rab11FIP1 KO, Rab11FIP2 KO, and heterozygous littermates. Rab11FIP1 KO mice showed normal fertility and body weight gain. More frequent lymphoid patches and infiltration of macrophages and neutrophils were identified in Rab11FIP1 KO mice before the development of rectal prolapse compared with control mice. The population of trefoil factor 3 (TFF3)-positive goblet cells was significantly lower, and the ratio of proliferative to nonproliferative cells was higher in Rab11FIP1 KO colons. Transcription signatures indicated that Rab11FIP1 deletion downregulated genes that mediate stress tolerance response, whereas genes mediating the response to infection were significantly upregulated, consistent with the inflammatory responses in the steady state. Lack of Rab11FIP1 also resulted in abnormal accumulation of subapical vesicles in colonocytes and the internalization of transmembrane mucin, MUC13, with Rab14. After DSS treatment, Rab11FIP1 KO mice showed greater body weight loss and more severe mucosal damage than those in heterozygous littermates. These findings suggest that Rab11FIP1 is important for cytoprotection mechanisms and for the maintenance of colonic mucosal integrity.NEW & NOTEWORTHY Although Rab11FIP1 is important in membrane trafficking in epithelial cells, the gastrointestinal phenotype of Rab11FIP1 knockout (KO) mice had never been reported. This study demonstrated that Rab11FIP1 loss induces mistrafficking of Rab14 and MUC13 and decreases in colonic goblet cells, resulting in impaired mucosal integrity.

Lysophosphatidic Acid Increases Maturation of Brush Borders and SGLT1 Activity in MYO5B-deficient Mice, a Model of Microvillus Inclusion Disease.

BACKGROUND & AIM: Myosin VB (MYO5B) is an essential trafficking protein for membrane recycling in gastrointestinal epithelial cells. The inactivating mutations of MYO5B cause the congenital diarrheal disease, microvillus inclusion disease (MVID). MYO5B deficiency in mice causes mislocalization of SGLT1 and NHE3, but retained apical function of CFTR, resulting in malabsorption and secretory diarrhea. Activation of lysophosphatidic acid (LPA) receptors can improve diarrhea, but the effect of LPA on MVID symptoms is unclear. We investigated whether LPA administration can reduce the epithelial deficits in MYO5B-knockout mice. METHODS: Studies were conducted with tamoxifen-induced, intestine-specific knockout of MYO5B (VilCreERT2;Myo5bflox/flox) and littermate controls. Mice were given LPA, an LPAR2 agonist (GRI977143), or vehicle for 4 days after a single injection of tamoxifen. Apical SGLT1 and CFTR activities were measured in Üssing chambers. Intestinal tissues were collected, and localization of membrane transporters was evaluated by immunofluorescence analysis in tissue sections and enteroids. RNA sequencing and enrichment analysis were performed with isolated jejunal epithelial cells. RESULTS: Daily administration of LPA reduced villus blunting, frequency of multivesicular bodies, and levels of cathepsins in intestinal tissues of MYO5B-knockout mice compared with vehicle administration. LPA partially restored the brush border height and the localization of SGLT1 and NHE3 in small intestine of MYO5B-knockout mice and enteroids. The SGLT1-dependent short-circuit current was increased and abnormal CFTR activities were decreased in jejunum from MYO5B-knockout mice given LPA compared with vehicle. CONCLUSIONS: LPA may regulate a MYO5B-independent trafficking mechanism and brush border maturation, and therefore be developed for treatment of MVID.

Loss of MYO5B Leads to Reductions in Na+ Absorption With Maintenance of CFTR-Dependent Cl- Secretion in Enterocytes.

BACKGROUND & AIMS: Inactivating mutations in MYO5B cause microvillus inclusion disease (MVID), but the physiological cause of the diarrhea associated with this disease is unclear. We investigated whether loss of MYO5B results in aberrant expression of apical enterocyte transporters. METHODS: We studied alterations in apical membrane transporters in MYO5B-knockout mice, as well as mice with tamoxifen-inducible, intestine-specific disruption of Myo5b (VilCreERT2;Myo5bflox/flox mice) or those not given tamoxifen (controls). Intestinal tissues were collected from mice and analyzed by immunostaining, immunoelectron microscopy, or cultured enteroids were derived. Functions of brush border transporters in intestinal mucosa were measured in Ussing chambers. We obtained duodenal biopsy specimens from individuals with MVID and individuals without MVID (controls) and compared transporter distribution by immunocytochemistry. RESULTS: Compared to intestinal tissues from littermate controls, intestinal tissues from MYO5B-knockout mice had decreased apical localization of SLC9A3 (also called NHE3), SLC5A1 (also called SGLT1), aquaporin (AQP) 7, and sucrase isomaltase, and subapical localization of intestinal alkaline phosphatase and CDC42. However, CFTR was present on apical membranes of enterocytes from MYO5B knockout and control mice. Intestinal biopsies from patients with MVID had subapical localization of NHE3, SGLT1, and AQP7, but maintained apical CFTR. After tamoxifen administration, VilCreERT2;Myo5bflox/flox mice lost apical NHE3, SGLT1, DRA, and AQP7, similar to germline MYO5B knockout mice. Intestinal tissues from VilCreERT2;Myo5bflox/flox mice had increased CFTR in crypts and CFTR localized to the apical membranes of enterocytes. Intestinal mucosa from VilCreERT2;Myo5bflox/flox mice given tamoxifen did not have an intestinal barrier defect, based on Ussing chamber analysis, but did have decreased SGLT1 activity and increased CFTR activity. CONCLUSIONS: Although trafficking of many apical transporters is regulated by MYO5B, trafficking of CFTR is largely independent of MYO5B. Decreased apical localization of NHE3, SGLT1, DRA, and AQP7 might be responsible for dysfunctional water absorption in enterocytes of patients with MVID. Maintenance of apical CFTR might exacerbate water loss by active secretion of chloride into the intestinal lumen.

Efficacy of self-management exercise program with spa therapy for behavioral management of knee osteoarthritis: research protocol for a quasi-randomized controlled trial (GEET one).

BACKGROUND: Osteoarthritis (OA) is not limited to joint pain and stiffness, which can lead to disability; it is also linked to comorbidities such as overweight, obesity and fears and beliefs related to the pathology. The knee OA population appears more affected by these risk factors and has a lower physical activity (PA) level than the general population. The key challenge for OA treatment is increasing the PA level to decrease the risk factors. METHODS: We aim to perform a prospective, multicentric, quasi-randomized controlled trial with an alternate-month design (1-month periods). People aged 50-75 years old with symptomatic knee OA (stage I-IV Kellgren and Lawrence scale) with low and moderate PA level will be included in 3 spa therapy resorts. The experimental arm will receive 5 self-management exercise sessions (1.5 h each; education, aerobics, strength training, range of motion) + an information booklet + 18 sessions (1 h each) of spa therapy treatment (STT). The active comparator arm will receive an information booklet + 18 sessions of STT. The primary outcome will be a change at 3 months in PA level (International Physical Activity Questionnaire short form score). Secondary outcomes will be function (WOMAC) pain (numerical scale), anxiety/depression (HAD), fears and beliefs about OA (KOFBeQ) and arthritis self-efficacy (ASES). The barriers to and facilitators of regular PA practice will be assessed by using specific items specifically designed for the study because of lack of any reference scale. DISCUSSION: The study could demonstrate the impact of a self-management exercise program associated with spa therapy in the medium term by increasing PA level in people with OA. A benefit for ameliorating fears and beliefs and anxiety/depression and improving self-efficacy will also be analysed. The findings could offer new prospects while establishing best clinical practice guidelines for this population. TRIAL REGISTRATION: ClinicalTrials.gov NCT02598804 (November 5, 2015).

Role of JunB in adenosine A2B receptor-mediated vascular endothelial growth factor production.

Interstitial adenosine stimulates neovascularization in part through A2B adenosine receptor-dependent upregulation of vascular endothelial growth factor (VEGF). In the current study, we tested the hypothesis that A2B receptors upregulate JunB, which can contribute to stimulation of VEGF production. Using the human microvascular endothelial cell line, human mast cell line, mouse cardiac Sca1-positive stromal cells, and mouse Lewis lung carcinoma (LLC) cells, we found that adenosine receptor-dependent upregulation of VEGF production was associated with an increase in VEGF transcription, activator protein-1 (AP-1) activity, and JunB accumulation in all cells investigated. Furthermore, the expression of JunB, but not the expression of other genes encoding transcription factors from the Jun family, was specifically upregulated. In LLC cells expressing A2A and A2B receptor transcripts, only the nonselective adenosine agonist NECA (5'-N-ethylcarboxamidoadenosine), but not the selective A2A receptor agonist CGS21680 [2-p-(2-carboxyethyl) phenylethylamino-5'-N-ethylcarboxamidoadenosine], significantly increased JunB reporter activity and JunB nuclear accumulation, which were inhibited by the A2B receptor antagonist PSB603 [(8-[4-[4-((4-chlorophenzyl)piperazide-1-sulfonyl)phenyl]]-1-propylxanthine]. Using activators and inhibitors of intracellular signaling, we demonstrated that A2B receptor-dependent accumulation of JunB protein and VEGF secretion share common intracellular pathways. NECA enhanced JunB binding to the murine VEGF promoter, whereas mutation of the high-affinity AP-1 site (-1093 to -1086) resulted in a loss of NECA-dependent VEGF reporter activity. Finally, NECA-dependent VEGF secretion and reporter activity were inhibited by the expression of a dominant negative JunB or by JunB knockdown. Thus, our data suggest an important role of the A2B receptor-dependent upregulation of JunB in VEGF production and possibly other AP-1-regulated events.

Comparison of self-management and spa therapy for upper-extremity musculoskeletal disorders: A randomized controlled trial.

BACKGROUND: Musculoskeletal disorders (MSDs) are common in the workplace and are a public health issue. Persistent pain despite conservative treatment or surgery may lead to poor long-term outcomes. OBJECTIVE: To compare the effect of a combined 6-day program of exercise, self-management workshops and spa therapy with self-management on functional capacity in personal and professional daily life at 3 months in people with musculoskeletal disorders. METHODS: We conducted a prospective, randomized controlled trial. Participants were employed (any type of work) and aged between 18 and 65 years, with latent or symptomatic upper extremity MSDs, with or without a history of sick leave. They were randomized to participate in 6 days (2 h per day) of spa therapy, exercise, and self-management workshops immediately (intervention) or at 3 months (control). The control group performed self-management until 3 months. The primary outcome was the score on the self-reported Quick Disability of Arm-Shoulder-Hand (QuickDASH) at 3 months. The primary analysis was conducted using analysis of covariance with baseline QuickDASH score as the covariate. RESULTS: In total, 150 participants were randomized (85 % women): 78 to the control group and 72 to the intervention group. At 3 months, the QuickDASH total and work scores did not differ between groups (effect-size [ES] = -0.15, 95 %CI, -0.38 to 0.09, p = 0.215, and ES = -0.11, 95 % CI, -0.35 to 0.12, p = 0.343). However, QuickDASH sport/performing arts score was significantly different between randomization groups at 3 months (ES =-0.25, 95 % CI, -0.48 to -0.02, p = 0.035). CONCLUSIONS: This study provided no evidence in favor of a short-course, personalized self-management, intensive spa therapy intervention over self-management alone for the management of upper-extremity MSDs. TRIAL REGISTRATION: ClinicalTrials.gov (NCT02702466) retrospectively registered.

Enhancing Prosthetic Vision by Upgrade of a Subretinal Photovoltaic Implant in situ.

In patients with atrophic age-related macular degeneration, subretinal photovoltaic implant (PRIMA) provided visual acuity up to 20/440, matching its 100µm pixels size. Next-generation implants with smaller pixels should significantly improve the acuity. This study in rats evaluates removal of a subretinal implant, replacement with a newer device, and the resulting grating acuity in-vivo. Six weeks after the initial implantation with planar and 3-dimensional devices, the retina was re-detached, and the devices were successfully removed. Histology demonstrated a preserved inner nuclear layer. Re-implantation of new devices into the same location demonstrated retinal re-attachment to a new implant. New devices with 22µm pixels increased the grating acuity from the 100µm capability of PRIMA implants to 28µm, reaching the limit of natural resolution in rats. Reimplanted devices exhibited the same stimulation threshold as for the first implantation of the same implants in a control group. This study demonstrates the feasibility of safely upgrading the subretinal photovoltaic implants to improve prosthetic visual acuity.

Adenosinergic regulation of the expansion and immunosuppressive activity of CD11b+Gr1+ cells.

Extracellular adenosine and purine nucleotides are elevated in many pathological situations associated with the expansion of CD11b(+)Gr1(+) myeloid-derived suppressor cells (MDSCs). Therefore, we tested whether adenosinergic pathways play a role in MDSC expansion and functions. We found that A(2B) adenosine receptors on hematopoietic cells play an important role in accumulation of intratumoral CD11b(+)Gr1(high) cells in a mouse Lewis lung carcinoma model in vivo and demonstrated that these receptors promote preferential expansion of the granulocytic CD11b(+)Gr1(high) subset of MDSCs in vitro. Flow cytometry analysis of MDSCs generated from mouse hematopoietic progenitor cells revealed that the CD11b(+)Gr-1(high) subset had the highest levels of CD73 (ecto-5'-nucleotidase) expression (Δmean fluorescence intensity [MFI] of 118.5 ± 16.8), followed by CD11b(+)Gr-1(int) (ΔMFI of 57.9 ± 6.8) and CD11b(+)Gr-1(-/low) (ΔMFI of 12.4 ± 1.0) subsets. Even lower levels of CD73 expression were found on Lewis lung carcinoma tumor cells (ΔMFI of 3.2 ± 0.2). The high levels of CD73 expression in granulocytic CD11b(+)Gr-1(high) cells correlated with high levels of ecto-5'-nucleotidase enzymatic activity. We further demonstrated that the ability of granulocytic MDSCs to suppress CD3/CD28-induced T cell proliferation was significantly facilitated in the presence of the ecto-5'-nucleotidase substrate 5'-AMP. We propose that generation of adenosine by CD73 expressed at high levels on granulocytic MDSCs may promote their expansion and facilitate their immunosuppressive activity.

Effectiveness of Ultrasound-guided VS Electrical-stimulation-guided Botulinum Toxin Injections in Triceps Surae Spasticity after Stroke: A Randomized Controlled Study.

OBJECTIVE: To compare the efficacy of botulinum toxin injections using ultrasound-guidance vs electrical-stimulation-guidance in triceps surae (soleus and gastrocnemius) spasticity after stroke. DESIGN: A clinical, single-centre, prospective, interventional, single-blind, cross-over, randomized trial, with outpatients in the tertiary care hospital. After randomization, subjects received electrical-stimulation-guided, followed by ultrasound-guided abobotulinumtoxinA injection (n = 15), or the same 2 procedures in the reverse order (n = 15) with the same operator, 4 months apart. The primary endpoint was the Tardieu scale with the knee straight at 1 month after injection. RESULTS: The 2 groups did not differ in Tardieu scale score (effect size = 0.15, 95% confidence interval (95% CI) -0.22 to 0.51, p = 0.43). In addition, the muscle localization technique used had no influence on walking speed, pain on injection or spasticity, assessed at 1 month after the injection, using the modified Ashworth scale. Ultrasound-guided injections were faster to administer than electrical-stimulation-guided injections. CONCLUSION: In agreement with previous research, no differences were found in the efficacy of ultrasound-guided or electrical-stimulation-guided abobotulinumtoxinA injections in triceps surae spasticity after stroke. Both techniques are of equal use in guiding muscle localization for botulinum toxin injections in spastic triceps surae.

Evaluation of the impact of a smartphone application on adherence to home exercise program for people with chronic low back pain: research protocol for a pilot randomised controlled trial.

INTRODUCTION: Multidisciplinary rehabilitation programmes are highly recommended for individuals with the most disabling low back pain (LBP). However, the long-term adherence to regular home exercise is often poor. We aim to perform a prospective, controlled, pilot, randomised study that will evaluate the impact of a smartphone application on adherence to exercise programme for people with chronic LBP (CLBP). METHODS AND ANALYSIS: 120 participants with non-specific CLBP aged 18-65 years will be recruited and randomised in two groups: an experimental group benefitting from education in the application's use in addition to a conventional multidisciplinary rehabilitation programme (exercises and self-management education) and a control group who will only participate in the multidisciplinary rehabilitation programme. Both groups will undergo the programme 5 days a week for 3 weeks. The primary outcome will be a change in patient's adherence to physical exercise (Exercise Adherence Rating Scale) at 6 months. Secondary outcomes will be function (Oswestry Disability Index), beliefs concerning physical activity (Evaluation of Physical Activity Perception), pain (Numeric Rating Scale), and physical capacity and qualitative adherence (video).Statistical analyses will be performed according to intention to treat. A linear mixed model will be used to compare the primary endpoint between groups at 6 months post-randomisation.The study could demonstrate the impact of using a smartphone application on adherence to exercise programme in people with CLBP. We hypothesise that the application's use will improve outcomes through improved exercise adherence. ETHICS AND DISSEMINATION: The study was approved by the medical ethics committee of Ile de France 3. The results of this study will be disseminated in peer-reviewed publications and presentations at international scientific meetings and will also be disseminated to the participants. TRIAL REGISTRATION NUMBER: NCT04264949.

Role of A2B adenosine receptors in regulation of paracrine functions of stem cell antigen 1-positive cardiac stromal cells.

The existence of multipotent cardiac stromal cells expressing stem cell antigen (Sca)-1 has been reported, and their proangiogenic properties have been demonstrated in myocardial infarction models. In this study, we tested the hypothesis that stimulation of adenosine receptors on cardiac Sca-1(+) cells up-regulates their secretion of proangiogenic factors. We found that Sca-1 is expressed in subsets of mouse cardiac stromal CD31(-) and endothelial CD31(+) cells. The population of Sca-1(+)CD31(+) endothelial cells was significantly reduced, whereas the population of Sca-1(+)CD31(-) stromal cells was increased 1 week after myocardial infarction, indicating their relative functional importance in this pathophysiological process. An increase in adenosine levels in adenosine deaminase-deficient mice in vivo significantly augmented vascular endothelial growth factor (VEGF) production in cardiac Sca-1(+)CD31(-) stromal cells but not in Sca-1(+)CD31(+) endothelial cells. We found that mouse cardiac Sca-1(+)CD31(-) stromal cells predominantly express mRNA encoding A(2B) adenosine receptors. Stimulation of adenosine receptors significantly increased interleukin (IL)-6, CXCL1 (a mouse ortholog of human IL-8), and VEGF release from these cells. Using conditionally immortalized Sca-1(+)CD31(-) stromal cells obtained from wild-type and A(2B) receptor knockout mouse hearts, we demonstrated that A(2B) receptors are essential for adenosine-dependent up-regulation of their paracrine functions. We found that the human heart also harbors a population of stromal cells similar to the mouse cardiac Sca-1(+)CD31(-) stromal cells that increase release of IL-6, IL-8, and VEGF in response to A(2B) receptor stimulation. Thus, our study identified A(2B) adenosine receptors on cardiac stromal cells as potential targets for up-regulation of proangiogenic factors in the ischemic heart.

Pixel size limit of the PRIMA implants: from humans to rodents and back.

Objective.Retinal prostheses aim at restoring sight in patients with retinal degeneration by electrically stimulating the inner retinal neurons. Clinical trials with patients blinded by atrophic age-related macular degeneration using the PRIMA subretinal implant, a 2 × 2 mm array of 100µm-wide photovoltaic pixels, have demonstrated a prosthetic visual acuity closely matching the pixel size. Further improvement in resolution requires smaller pixels, which, with the current bipolar design, necessitates more intense stimulation.Approach.We examine the lower limit of the pixel size for PRIMA implants by modeling the electric field, leveraging the clinical benchmarks, and using animal data to assess the stimulation strength and contrast of various patterns. Visually evoked potentials measured in Royal College of Surgeons rats with photovoltaic implants composed of 100µm and 75µm pixels were compared to clinical thresholds with 100µm pixels. Electrical stimulation model calibrated by the clinical and rodent data was used to predict the performance of the implant with smaller pixels.Main results.PRIMA implants with 75µm bipolar pixels under the maximum safe near-infrared (880 nm) illumination of 8 mW mm-2with 30% duty cycle (10 ms pulses at 30 Hz) should provide a similar perceptual brightness as with 100µm pixels under 3 mW mm-2irradiance, used in the current clinical trials. Contrast of the Landolt C pattern scaled down to 75µm pixels is also similar under such illumination to that with 100µm pixels, increasing the maximum acuity from 20/420 to 20/315.Significance.Computational modeling defines the minimum pixel size of the PRIMA implants as 75µm. Increasing the implant width from 2 to 3 mm and reducing the pixel size from 100 to 75µm will nearly quadrupole the number of pixels, which should be very beneficial for patients. Smaller pixels of the same bipolar flat geometry would require excessively intense illumination, and therefore a different pixel design should be considered for further improvement in resolution.

Photovoltaic implant simulator reveals resolution limits in subretinal prosthesis.

Objective.PRIMA, the photovoltaic subretinal prosthesis, restores central vision in patients blinded by atrophic age-related macular degeneration (AMD), with a resolution closely matching the 100µm pixel size of the implant. Improvement in resolution requires smaller pixels, but the resultant electric field may not provide sufficient stimulation strength in the inner nuclear layer (INL) or may lead to excessive crosstalk between neighboring electrodes, resulting in low contrast stimulation patterns. We study the approaches to electric field shaping in the retina for prosthetic vision with higher resolution and improved contrast.Approach.We present a new computational framework, Retinal Prosthesis Simulator (RPSim), that efficiently computes the electric field in the retina generated by a photovoltaic implant with thousands of electrodes. Leveraging the PRIMA clinical results as a benchmark, we use RPSim to predict the stimulus strength and contrast of the electric field in the retina with various pixel designs and stimulation patterns.Main results.We demonstrate that by utilizing monopolar pixels as both anodes and cathodes to suppress crosstalk, most patients may achieve resolution no worse than 48µm. Closer proximity between the electrodes and the INL, achieved with pillar electrodes, enhances the stimulus strength and contrast and may enable 24µm resolution with 20µm pixels, at least in some patients.Significance.A resolution of 24µm on the retina corresponds to a visual acuity of 20/100, which is over 4 times higher than the current best prosthetic acuity of 20/438, promising a significant improvement of central vision for many AMD patients.

Identification of Responders to Balneotherapy among Adults over 60 Years of Age with Chronic Low Back Pain: A Pilot Study with Trajectory Model Analysis.

Balneotherapy may be a relevant treatment for chronic low back pain (LBP) in individuals > 60 years old. This pilot study aimed to determine the effectiveness of balneotherapy for chronic LBP in people > 60 years old and to determine profiles of responders with trajectory model analysis. This was a pilot prospective open cohort study, with repeated measurements using validated questionnaires; participants were their own controls. The primary endpoint was the proportion of participants with a change in pain intensity between the start of treatment and 3 months after treatment assessed with a numeric scale (NS) from 0 to 100 mm, with an effect size (ES) > 0.5. The assessments involved questionnaires that were self-administered on days (D) 1 and 21 and at months 3 and 6. The secondary objective was to determine the profile of responders to balneotherapy. We included 78 patients (69.2% women), mean age 68.3 ± 5.3 years. The mean pain score on the NS was 48.8 ± 19.9 at D1 and 39.1 ± 20.5 at 3 months (p < 0.001). The ES was 0.47 [95% confidence interval [CI] 0.25 to 0.69] for the whole sample; 36% (28/78) had an ES > 0.5; 23% (18/78) had a moderate ES (0 to 0.5); and 41% (32/78) had an ES of zero (14/78) or < 0 (18/78), corresponding to increased pain intensity. The pain trajectory model showed that the change in pain between D1 and D21 for trajectory A (larger reduction in pain intensity) was -50% [95% CI -60 to -27], and for trajectory B (smaller reduction in pain intensity), it was -13% [-33 to 0] (p < 0.001). Between Day 1 and month 3, the change for trajectory A was -33% [-54; 0] and for trajectory B was -13% [-40 to 0] (p = 0.14). Finally, between D1 and month 6, the change for trajectory A was -50% [-60 to 0] and for trajectory B was -6% [-33 to 17] (p = 0.007). The patients in trajectory A reported performing more physical activity than those in trajectory B (p = 0.04). They were also less disabled, with a mean Oswestry Disability Index of 40.4 versus 45.7 for those in trajectory A and B, respectively, (p = 0.03) and had a higher total Arthritis Self-Efficacy Scale score. This real-life study of the effectiveness of balneotherapy on chronic LBP identified distinct pain trajectories and predictive variables for responders. These criteria could be used in decision-making regarding the prescription of balneotherapy, to ensure personalized management of chronic LBP.

Cell differentiation is disrupted by MYO5B loss through Wnt/Notch imbalance.

Functional loss of myosin Vb (MYO5B) induces a variety of deficits in intestinal epithelial cell function and causes a congenital diarrheal disorder, microvillus inclusion disease (MVID). The impact of MYO5B loss on differentiated cell lineage choice has not been investigated. We quantified the populations of differentiated epithelial cells in tamoxifen-induced, epithelial cell-specific MYO5B-knockout (VilCreERT2 Myo5bfl/fl) mice utilizing digital image analysis. Consistent with our RNA-sequencing data, MYO5B loss induced a reduction in tuft cells in vivo and in organoid cultures. Paneth cells were significantly increased by MYO5B deficiency along with expansion of the progenitor cell zone. We further investigated the effect of lysophosphatidic acid (LPA) signaling on epithelial cell differentiation. Intraperitoneal LPA significantly increased tuft cell populations in both control and MYO5B-knockout mice. Transcripts for Wnt ligands were significantly downregulated by MYO5B loss in intestinal epithelial cells, whereas Notch signaling molecules were unchanged. Additionally, treatment with the Notch inhibitor dibenzazepine (DBZ) restored the populations of secretory cells, suggesting that the Notch pathway is maintained in MYO5B-deficient intestine. MYO5B loss likely impairs progenitor cell differentiation in the small intestine in vivo and in vitro, partially mediated by Wnt/Notch imbalance. Notch inhibition and/or LPA treatment may represent an effective therapeutic approach for treatment of MVID.

Recruitment of Polarity Complexes and Tight Junction Proteins to the Site of Apical Bulk Endocytosis.

BACKGROUND & AIMS: The molecular motor, Myosin Vb (MYO5B), is well documented for its role in trafficking cargo to the apical membrane of epithelial cells. Despite its involvement in regulating apical proteins, the role of MYO5B in cell polarity is less clear. Inactivating mutations in MYO5B result in microvillus inclusion disease (MVID), a disorder characterized by loss of key apical transporters and the presence of intracellular inclusions in enterocytes. We previously identified that inclusions in Myo5b knockout (KO) mice form from invagination of the apical brush border via apical bulk endocytosis. Herein, we sought to elucidate the role of polarity complexes and tight junction proteins during the formation of inclusions. METHODS: Intestinal tissue from neonatal control and Myo5b KO littermates was analyzed by immunofluorescence to determine the localization of polarity complexes and tight junction proteins. RESULTS: Proteins that make up the apical polarity complexes-Crumbs3 and Pars complexes-were associated with inclusions in Myo5b KO mice. In addition, tight junction proteins were observed to be concentrated over inclusions that were present at the apical membrane of Myo5b-deficient enterocytes in vivo and in vitro. Our mouse findings are complemented by immunostaining in a large animal swine model of MVID genetically engineered to express a human MVID-associated mutation that shows an accumulation of Claudin-2 over forming inclusions. The findings from our swine model of MVID suggest that a similar mechanism of tight junction accumulation occurs in patients with MVID. CONCLUSIONS: These data show that apical bulk endocytosis involves the altered localization of apical polarity proteins and tight junction proteins after loss of Myo5b.

Attenuation of chronic pulmonary inflammation in A2B adenosine receptor knockout mice.

Pharmacologic evidence suggests that activation of A(2B) adenosine receptors results in proinflammatory effects relevant to the progression of asthma, a chronic lung disease associated with elevated interstitial adenosine concentrations in the lung. This concept has been challenged by the finding that genetic removal of A(2B) receptors leads to exaggerated responses in models of acute inflammation. Therefore, the goal of our study was to determine the effects of A(2B) receptor gene ablation in the context of ovalbumin-induced chronic pulmonary inflammation. We found that repetitive airway allergen challenge induced a significant increase in adenosine levels in fluid recovered by bronchoalveolar lavage. Genetic ablation of A(2B) receptors significantly attenuated allergen-induced chronic pulmonary inflammation, as evidenced by a reduction in the number of bronchoalveolar lavage eosinophils and in peribronchial eosinophilic infiltration. The most striking difference in the pulmonary inflammation induced in A(2B) receptor knockout (A(2B)KO) and wild-type mice was the lack of allergen-induced IL-4 release in the airways of A(2B)KO animals, in line with a significant reduction in IL-4 protein and mRNA levels in lung tissue. In addition, attenuation of allergen-induced transforming growth factor-beta release in airways of A(2B)KO mice correlated with reduced airway smooth muscle and goblet cell hyperplasia/hypertrophy. In conclusion, genetic removal of A(2B) adenosine receptors in mice leads to inhibition of allergen-induced chronic pulmonary inflammation and airway remodeling. These findings are in agreement with previous pharmacologic studies suggesting a deleterious role for A(2B) receptor signaling in chronic lung inflammation.

Adenosine receptors in regulation of dendritic cell differentiation and function.

Differentiation of functional dendritic cells (DCs) critically depends on the microenvironment. DCs differentiate in hypoxic tumor sites and inflamed or damaged tissue. Because local concentrations of adenosine reach high physiologically relevant levels in these conditions, we assessed the expression of adenosine receptors and the effect of their activation on differentiation of human monocytes and mouse peritoneal macrophages and hematopoietic progenitor cells (HPCs) into myeloid DCs. Stimulation of adenosine receptors skews DC differentiation toward a distinct cell population characterized by expression of both DC and monocyte/macrophage cell surface markers. Pharmacologic analysis and experiments with cells from A(2B) adenosine receptor knockout mice identified A(2B) receptor as the mediator of adenosine effects on DCs. Unlike normal myeloid DCs, adenosine-differentiated DCs have impaired allostimulatory activity and express high levels of angiogenic, pro-inflammatory, immune suppressor, and tolerogenic factors, including VEGF, IL-8, IL-6, IL-10, COX-2, TGF-beta, and IDO. They promoted tumor growth if injected into tumors implanted in mice. Using adenosine desaminase knockout animals, we showed that DCs with proangiogenic phenotype are highly abundant under conditions associated with elevated levels of extracellular adenosine in vivo. Adenosine signaling through A(2B) receptor is an important factor of aberrant DC differentiation and generation of tolerogenic, angiogenic, and proinflammatory cells.