Piezo1-Regulated Mechanotransduction Controls Flow-Activated Lymphatic Expansion.

BACKGROUND: Mutations in PIEZO1 (Piezo type mechanosensitive ion channel component 1) cause human lymphatic malformations. We have previously uncovered an ORAI1 (ORAI calcium release-activated calcium modulator 1)-mediated mechanotransduction pathway that triggers lymphatic sprouting through Notch downregulation in response to fluid flow. However, the identity of its upstream mechanosensor remains unknown. This study aimed to identify and characterize the molecular sensor that translates the flow-mediated external signal to the Orai1-regulated lymphatic expansion. METHODS: Various mutant mouse models, cellular, biochemical, and molecular biology tools, and a mouse tail lymphedema model were employed to elucidate the role of Piezo1 in flow-induced lymphatic growth and regeneration. RESULTS: Piezo1 was found to be abundantly expressed in lymphatic endothelial cells. Piezo1 knockdown in cultured lymphatic endothelial cells inhibited the laminar flow-induced calcium influx and abrogated the flow-mediated regulation of the Orai1 downstream genes, such as KLF2 (Krüppel-like factor 2), DTX1 (Deltex E3 ubiquitin ligase 1), DTX3L (Deltex E3 ubiquitin ligase 3L,) and NOTCH1 (Notch receptor 1), which are involved in lymphatic sprouting. Conversely, stimulation of Piezo1 activated the Orai1-regulated mechanotransduction in the absence of fluid flow. Piezo1-mediated mechanotransduction was significantly blocked by Orai1 inhibition, establishing the epistatic relationship between Piezo1 and Orai1. Lymphatic-specific conditional Piezo1 knockout largely phenocopied sprouting defects shown in Orai1- or Klf2- knockout lymphatics during embryo development. Postnatal deletion of Piezo1 induced lymphatic regression in adults. Ectopic Dtx3L expression rescued the lymphatic defects caused by Piezo1 knockout, affirming that the Piezo1 promotes lymphatic sprouting through Notch downregulation. Consistently, transgenic Piezo1 expression or pharmacological Piezo1 activation enhanced lymphatic sprouting. Finally, we assessed a potential therapeutic value of Piezo1 activation in lymphatic regeneration and found that a Piezo1 agonist, Yoda1, effectively suppressed postsurgical lymphedema development. CONCLUSIONS: Piezo1 is an upstream mechanosensor for the lymphatic mechanotransduction pathway and regulates lymphatic growth in response to external physical stimuli. Piezo1 activation presents a novel therapeutic opportunity for preventing postsurgical lymphedema. The Piezo1-regulated lymphangiogenesis mechanism offers a molecular basis for Piezo1-associated lymphatic malformation in humans.

Novel Endoscopic Polypectomy Surveillance Technique for Fundic Gland Polyps in Familial Adenomatous Polyposis Can Improve Early Detection of Dysplasia and Gastric Cancer.

INTRODUCTION: Fundic gland polyps (FGPs) are commonly found in patients with familial adenomatous polyposis (FAP) and are considered benign. Biopsies are not routinely performed, and conventional forceps may be time-consuming and/or yield nonrepresentative histology. The purpose of this study was to evaluate the role of a novel endoscopic polypectomy surveillance (EPS), a large volume cold-snare polypectomy technique of random FGPs, in the incidence of dysplasia and gastric cancer (GC) in FAP. METHODS: This is a retrospective longitudinal cohort of patients with FAP referred to a tertiary care center for duodenal adenoma surveillance and who underwent EPS of FGPs between 2001 and 2019. Demographic, endoscopic, and clinicopathologic information was reviewed. RESULTS: Thirty-five patients with FAP were identified at initial endoscopy by the mean age of 43.4 years (±12.8). One hundred thirteen surveillance endoscopies were performed in total using EPS. Dysplasia of FGPs was present on initial esophagogastroduodenoscopy in 7 patients (20%), and 13 additional patients (46.4%) progressed to low-grade dysplasia. Three patients (15%) who subsequently had progression to GC were found to have signet ring cell cancer within the foci of FGPs through EPS. One patient presented as metastatic GC. Progression from nondysplastic FGP to low-grade dysplasia occurred over 63 months (±46.3) with further progression to GC over 34 months (±8.5). Endoscopic risk factors for cancer were polyps >10 mm in size ( P < 0.001) and carpeting of polyps ( P < 0.001). The 5-year cumulative incidence of developing dysplasia was 35.7%. DISCUSSION: We identified that the incidence of dysplasia and GC is higher than previously reported in patients with FAP. Our study used a novel EPS technique and was able to identify GC within the foci of FGPs. Upper endoscopic guidelines should include a more rigorous sampling method for FGPs, such as EPS, to optimize early detection of dysplasia and GC.

4-(Hydroxymethyl)catechol Extracted From Fungi in Marine Sponges Attenuates Rheumatoid Arthritis by Inhibiting PI3K/Akt/NF-κB Signaling.

Rheumatoid arthritis (RA) is a progressive autoimmune disease specific to synovial joints; it causes joint damage and other systemic abnormalities, thereby leading to physical disability and early mortality. Marine sponge-derived fungi, Pestalotiopsis sp., secrete immunosuppressive compounds in the culture broth. In the present study, we isolated 4-(hydroxymethyl)catechol (4-HMC) from these fungal species, and evaluated its anti-RA effects using a murine collagen-induced arthritis model and tumor necrosis factor-α-stimulated human RA synovial fibroblasts. Oral 4-HMC administration decreased the clinical arthritis score, paw thickness, histologic and radiologic changes, and serum IgG1 and IgG2a levels. It prevented the proliferation of helper T (Th) 1/Th17 CD4+ lymphocytes isolated from inguinal lymph nodes, thereby reducing inflammatory cytokine production in CIA mice. It decreased the expression of inflammatory mediators, including cytokines and matrix metalloproteinases (MMPs), both in vitro and in vivo. We observed that 4-HMC suppresses Th immune responses and MMP expression to inhibit inflammatory cytokine production in human RA synovial fibroblasts by modulating the PI3K/Akt/NF-κB pathway. These results verify the anti-RA potential of 4-HMC.

Differences in Patient-Reported Outcomes Between Unicompartmental and Total Knee Arthroplasties: A Propensity Score-Matched Analysis.

BACKGROUND: The purpose of this study was to compare the patient-reported outcomes regarding joint awareness, function, and satisfaction after unicompartmental knee arthroplasty (UKA) and total knee arthroplasty (TKA). METHODS: We identified all patients who underwent a UKA or TKA at our institution between September 2011 and March 2014, with a minimum follow-up of 2 years. Propensity score matching was performed for age, gender, body mass index, operation side, and the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) score. One hundred UKAs to 100 TKAs were matched. Each knee was evaluated according to the WOMAC score, Forgotten Joint Score (FJS), High Flexion Knee Score (HFKS) and patient's satisfaction at postoperative 2 years. RESULTS: There was no significant difference in WOMAC score at postoperative 2 years between UKA and TKA groups. However, the FJS of the UKA group was significantly higher than that of the TKA group (67.3 ± 19.8 and 60.6 ± 16.6, respectively; P = .011). The HFKS was also significantly higher in the UKA group compared with the TKA group (34.4 ± 6.4 and 31.3 ± 5.2, respectively; P < .001). Eighty-six percent of all patients who underwent UKA were satisfied compared with 71% of those who underwent TKA (P = .027). CONCLUSION: Patients who underwent UKA had higher FJS, HFKS, and satisfaction rate when compared with patients who underwent TKA, indicating that UKA facilitated less knee awareness and better function and satisfaction than TKA.

Cerebral aqueduct block attenuates cardio-renal injuries in post-DOCA-NaCl-hypertensive Dahl R rats.

The systemic and/or local effects of the hydrocephalic brain were investigated in DOCA-NaCl-hypertensive Dahl R rats induced by 250 mg kg(-1) DOCA in silicone and 1% saline water. After a 1-week recovery with 0.3% NaCl chow and tap water, one group had the aqueduct of Sylvius blocked with silicone and epoxy materials with a control sham group matching mean blood pressure (BP) and body weight. The 4-week-postsurgery BP on the 0.3% NaCl diet averaged 161±3.2 in the sham group and 146±2.3 mm Hg in the blocked group (P<0.0001). Both groups were then given an 8% NaCl diet and after 4 weeks, the sham group's BP was increased further with markedly increased mortality: 186 mm Hg vs. 154 mm Hg (P<0.0001); 12 sham rats died after 11 weeks, while all the blocked rats survived (P<0.0001). A transient change in plasma Na levels was observed in the blocked group after 48 h on the 8% NaCl diet. At 14 weeks, 0 sham rats survived, compared with 10 out of 16 blocked rats (P<0.0001). After 11 weeks on 8% NaCl, the average tail venous pressure in the sham group was significantly higher than that of the blocked rats (P<0.0001) indicating the end stage of renal and heart failure. The hearts and kidneys weighed significantly more in the sham vs. the blocked rats (P<0.0001 for both groups). These results indicate that the aqueduct block prevents post-DOCA hypertension and cardio-renal injuries, suggesting that centralized third ventricular brain signaling has a role in salt-genetic hypertension.

Wistar-Kyoto and spontaneously hypertensive rat blood pressure after embryo transfer into different wombs and cross-suckling.

Blood pressure (BP) varies based on genetic and environmental factors. To test genetic and environmental influences on body weight (BW) and BP, one-cell homozygous embryos were transferred into spontaneously hypertensive (SHR, pup:shr) or (Wistar-Kyoto normotensive [WKY], pup:wky) normotensive rats' oviducts (embryos: s,w; oviduct-uterine: S,W), cross-suckled at birth (nurses S,W) and weaned to normal diets at day-21. BP at day-120 was measured by radiotelemetry and analyzed by methods of linear least square rhythmometry and analysis of variance. Genetics dominantly affected shr BP, causing it to be significantly higher at birth (24.6 ± 1.8 in sS versus 21.8 ± 1.7 mmHg in wW, P < 0.005), and at day-120 (198 ± 0.5 in sSS versus 127 ± 0.2 mmHg in wWW, P < 0.001), with lower BW than those of wky (5.3 ± 0.2 versus 5.7 ± 0.2 g at birth, 332 ± 5 versus 404 ± 6 g at day-120, both P < 0.001). Surprisingly, uterine-suckling milieus lowered shr BP significantly at day-120 (198 ± 0.5 in sSS versus 178 ± 0.5, 147 ± 0.6, 179 ± 0.5 mmHg in sSW, sWS, sWW, respectively, all P < 0.01). BP was slightly elevated when wky-genetics were implanted into the S-uterine by 4 mmHg (wSW, P < 0.05), whereas implanting shr embryos into the W-uterine environment (sWS) lowered BP by 51 mmHg (P < 0.001). In summary, the hypertensive shr-strain showed significantly lower BP when provided with an WKY-uterine environment and/or by WKY-nursing mothers, indicating that environment can modify genetic influences; yet the shr MESORs (rhythm-adjusted 24-h mean: midline estimating statistic of rhythm) lowered by WKY environments remained above MESORs encountered in wky-donors.

Heritability of erythrocyte sodium permeability: a possible genetic marker for hypertension.

Individuals genetically susceptible to hypertension may have preexisting membrane defects influencing cell sodium permeability. Fourteen Minnesotan families of Northern European descent were selected as having one or both progenitors with either high (HP) or low (LP) erythrocyte sodium permeability. We earlier found that over one-half of the (22)Na(+) influx into HP erythrocytes can be inhibited by micromolar amounts of furosemide, which has no apparent effect on LP erythrocytes. In these families, we find a significant midpoint parent/offspring correlation in the furosemide-sensitive component of erythrocyte (22)Na(+) flux rates (p <0.001). The relationship between parents and children in this metric trait is most consistent with a single locus, 2-allele system with variable expression, which suggests that enhanced furosemide-sensitive (22)Na(+) influx may be inherited as an autosomal recessive trait. Individuals with HP erythrocytes tend to have increased blood pressure and/or a family history of hypertension. The results were confirmed in these family samples with HP and LP (22)Na(+) influx (mmol/L RBC/hr): 0.404 +/- 0.03 vs 0.232 +/- 0.01 (p < 0.001); systolic blood pressure (mm Hg): 136 +/- 4 vs 108 +/- 4 (p < 0.001); and diastolic blood pressure (mm Hg): 89 +/- 2 vs 69 +/- 2 (p <0.001). These results may help to identify inherited factors involved in salt sensitive hypertension.

Predictive factors for lymph node metastasis and endoscopic treatment strategies for undifferentiated early gastric cancer.

BACKGROUND AND AIM: Although more than 80% of undifferentiated early gastric cancers (EGC) are not associated with lymph node metastasis, endoscopic mucosal resection is not generally accepted as a means of curative treatment because of an abundance of conflicting data concerning clinicopathological characteristics and prognoses. The aim of this study was to define a subgroup of undifferentiated EGC that could be cured by endoscopic treatment without the risk of lymph node metastasis. METHOD: A total of 591 patients surgically resected for undifferentiated EGC between January 1999 and March 2005 were reviewed. Associations between various clinicopathological factors and the presence of lymph node metastasis were analyzed to identify the risk factors of lymph node metastasis. RESULTS: Lymph node metastasis was found in 79 patients (13.4%). By multivariate logistic regression analysis, a tumor diameter 2.5 cm or larger, invasion into the middle third of the submucosal layer or deeper, and lymphatic involvement were identified as independent risk factors of lymph node metastasis (P < 0.001, respectively). Lymph node metastasis was not found in any patient with undifferentiated EGC smaller than 2.5 cm confined to the mucosa or upper third of the submucosal layer without lymphatic involvement. CONCLUSIONS: Undifferentiated intramucosal EGC smaller than 2.5 cm without lymphatic involvement was not associated with lymph node metastasis. Thus, we propose in this circumstance that endoscopic mucosal resection could be considered a definitive treatment without compromising the possibility of cure.

Prevention of erythropoietin-associated hypertension.

Hypertension is the most significant complication from treatment with erythropoietin (Epo). Can Epo-induced hypertension be eliminated? We examined systemic and local effects of our genetically engineered products, Epo-binding protein (Epo-bp) and anti-Epo-bp antibodies, on randomly assigned Sprague-Dawley rats at midnight, 4 am, 8 am, noon, 4 pm, and 8 pm. Blood pressure, hematocrit, and body weight were measured immediately before and after the completion of a 4-week, twice-weekly course of Epo (50 U/kg), Epo-bp, anti-Epo-bp antibodies, or physiological saline injections. Epo treatment increased hematocrit markedly overall as compared with the saline, Epo-bp, and anti-Epo-bp antibody groups (0.616 versus 0.427, 0.439, and 0.441, respectively) and at each of the 6 test times (all P<0.0001). Epo-bp and anti-Epo-bp antibody treatment with Epo had almost no effect on the Epo-induced hematocrit increase (0.616 versus 0.580 or 0.591, respectively). Circadian blood pressures for Epo versus saline, Epo-bp, and anti-Epo-bp antibody groups were 136.2+/-2.3 versus 116.2+/-1.7, 118.4+/-2.1, and 116.6+/-2.1 mm Hg, respectively (each P<0.0001). Significantly increased blood pressure was detected at noon, 4 pm, 8 pm, and midnight in Epo treatment. When Epo was given with Epo-bp or anti-Epo-bp antibodies, blood pressure was maintained at similar levels as in saline treatment (each P<0.0001) as compared with Epo treatment alone. Overall, body, brain, and heart weights were significantly lower in Epo treatment than those of other groups. Thus, Epo-bp and anti-Epo-bp antibodies eliminate Epo-induced hypertension without affecting hematocrit and blood volume.

Purification of biologically active human erythropoietin-binding protein and detection of its binding sites.

To purify human erythropoietin-binding protein (Epo-bp), the recombinant vector JYL26 was constructed by inserting human Epo-receptor cDNA by PCR into a pGEX-2T plasmid vector with a thrombin cleavage site. EpoRex-th fusion protein, containing glutathione-S-transferase (GST) and Epo-bp, was purified by glutathione-affinity chromatography. Biologically active pure human Epo-bp (MW=29 kDa) was then purified by Epo-agarose chromatography after cleaving off the GST by thrombin. Purified Epo-bp has a strong binding affinity to 125I-Epo, and unlabeled Epo eliminates the binding (p<0.0001). Trypsin digested Epo-bp to approximately 20 kDa and completely eliminated the binding. Thus, Epo-bp ligand binding is specific and it may require an intact Epo-bp. Ligand-binding sites were detected using fluorescein-labeling in our new products, Epo-bp and anti-Epo-bp antibody (alpha Epo-bp), in various blood cell progenitors, including megakaryocytes, erythroblasts, normoblasts, and myeloblasts, while fluorescein-labeled pre-immune Fab-treated cells did not show any binding. Epo, Epo-bp, and their antibodies were measured in serum and plasma specimens by enzyme immunoassay methods developed in our laboratory; Epo in serum and plasma: 25.4+/-2.17 and 24.2+/-2.35; Epo-bp in serum and plasma: 24.2+/-1.84 and 25.0+/-1.26 mU/ml, respectively. These assays are simple, sensitive, and precise, compared to the conventional Epo radioimmunoassay, and are more environmentally friendly than assays that use radioactive reagents.

About 7-day (circaseptan) and circadian changes in cold pressor test (CPT).

We assess circadian (CD) and circaseptan (CS) changes in the blood pressure (BP) response to a 1-min immersion of the hand into ice water, the cold pressor test (CPT). An about 8 mmHg BP increase in the CPT reported by others for health and an elevation >25 mmHg for patients with "hypertension" have been viewed as predisease, but variables related to the vascular system exhibit prominent CD, CS and circannuals, among other components of their genetically anchored time structure or chronome, and may also have to be considered. Hence, a 16-year-old adolescent (ML) immersed her hand into ice water every 2-4 h for 2 days and once daily in the morning thereafter for 1 week (N = 25), as did, only for <2 days (> or =24 h), four adults 20-66 years of age. BP and heart rate (HR) were monitored automatically around the clock at 15-min intervals and at 1-min intervals before and immediately after the CPT. Data were analyzed by cosinor. The difference between the first post-CPT BP value and the mean of the last seven values prior to the CPT was a measure of CPT response (at 1-min). Overall, ML's systolic (S) BP increased by 8.2 +/- 1.4 mmHg and her diastolic (D) BP by 6.2 +/- 0.9 mmHg (P < 0.001). Increases were found in 96% of the tests for SBP, 92% for DBP and 20% for HR. There was only one tie for HR (4%), resulting in a HR decrease in 76% of CPTs. The BP response to CPT at 1 min was CS-dependent. The CS double amplitudes are 12.5 +/- 4.1 for SBP (P = 0.019) and 7.8 +/- 2.8 mmHg for DBP (P = 0.030), with acrophases occurring on late Sunday, early Monday (at -50 degrees and -67 degrees from 00:00 h from Saturday to Sunday for SBP and DBP, respectively, with 360 degrees identical with 1 week). The response of HR did not allow the detection of a CS rhythm (P = 0.969). The CD response peaked in the early morning hours; with 24 h identical with 360 degrees and 0 degrees = local midnight, the acrophase (phi) for SBP is at -80 degrees and for DBP at -113 degrees, in keeping with earlier results from four adult subjects (SBP: -37 degrees; DBP: -42 degrees ), individual differences notwithstanding. The average timing of the largest overall response of BP to CPT coincides with the timing of the response to other stimuli. The CS acrophases coincide with the times of increased morbidity/mortality from vascular diseases. Chronomes, time structures broader than circadian, notably their about-weekly components, should be considered, not only at the extremes of life when the CS and about-half-weekly rhythms are particularly prominent in BP and HR but also in interpreting BP responses as a gauge of vascular disease status in adolescents.

Effects of diaphragmatic breathing on ambulatory blood pressure and heart rate.

Psychological procedures, such as self-hypnosis in the form of autogenic training, have been proposed for correcting a deviant, e.g., high blood pressure (BP). In view of the overwhelming evidence for the circadian (CD) stage dependence of any treatment effects, the CD stage dependence of the effects of diaphragmatic breathing (DB) on BP and heart rate (HR) was explored in data from a clinically healthy normotensive subject who, following 3 weeks of ambulatory monitoring as a reference standard, measured BP and HR with a manual monitor at 1-min intervals for 5 min before and after DB (three deep diaphragmatic breaths) and who performed DB for about 2 weeks at about 2-h intervals while awake. The 3-week data series were analyzed by cosinor, involving the least-squares fit of cosine curves with periods of 24, 12 and 168 h. A CD rhythm was detected for BP and HR (P < 0.001 in each case), peaking in the afternoon. Some about-weekly (circaseptan; CS) BP rhythms and 12-h (circasemidian) components were also statistically significant. DB was found to reduce systolic (S) BP. Overall, SBP decreased by 5.9 +/- 0.8 mmHg (P < 0.001) and diastolic (D) BP by 1.4 +/- 0.8 mmHg (P < 0.005), while HR remained at about the same average. The effect of DB on BP was CD-dependent, the largest response occurring in the afternoon, 2-3 h before the peaks in SBP and DBP found in the reference data of the same subject. There was also a 5-10% decrease in SBP around the weekend (Friday, Saturday and Sunday). The results on a single subject suggest the need to collect similar data on others for optimizing by clock-hour, day of the week, and eventually by the marker rhythms BP and/or HR the best times for DB and other procedures. The personalized best time for people on different work/rest schedules for relaxation may be several hours before their BP has reached its highest point in the 24-h span. HR may serve as a marker for DB timing, but the effect on HR of DB was only of borderline statistical significance in the subject investigated.

Protective effects in ambulatory blood pressure and centralized injuries in hydrocephalic Dahl rats on high and low NaCl diets.

Our previous studies in hydrocephalic Dahl rats indicated that the NaCl signal is perceived in the structures around the third brain ventricle. In the present study, we assessed 24-h ambulatory blood pressures (ABPs), weight (WT) gains, and the kidney and brain injuries in induced hydrocephalic and control group Dahl S rats on 0.3% and 6% NaCl diets. Constant WT gains were observed among low NaCl fed and high NaCl fed aqueduct blocked groups, whereas high NaCl-fed sham group rats were losing WT and dying off after 8 weeks. A circadian BP curve in the high NaCl sham group was very distinct from that in the other three groups with markedly increased peak pressure during the light cycle with very high amplitude as compared with all the other three groups (both P <.0001). There was not only a very unstable fluctuating curve present, but a day-night rhythm shift was also prominent in the high NaCl sham group. In a kidney cross-section, 54 rats on a 0.3% low NaCl diet for 14 weeks averaged 156 glomeruli. A total of 34 high NaCl fed, aqueduct blocked rats averaged 141 glomeruli, whereas 23 sham blocked rats averaged 102 glomeruli (-28%, P <.0001). A brain cross-section showed many small lacunae in high NaCl-fed sham rats as compared with the truly blocked rats (193 +/- 14 v 77 +/- 10 lacunae, a 2.5-fold reduction, P <.0001). The changes in the blocked group may be due to volume and electrolyte rebalance with reduced pressures in the brain volume-controlling center.