Planar cell polarity-dependent asymmetric organization of microtubules for polarized positioning of the basal body in node cells.

For left-right symmetry breaking in the mouse embryo, the basal body must become positioned at the posterior side of node cells, but the precise mechanism for this has remained unknown. Here, we examined the role of microtubules (MTs) and actomyosin in this basal body positioning. Exposure of mouse embryos to agents that stabilize or destabilize MTs or F-actin impaired such positioning. Active myosin II was detected at the anterior side of node cells before the posterior shift of the basal body, and this asymmetric activation was lost in Prickle and dachsous mutant embryos. The organization of basal-body associated MTs (baMTs) was asymmetric between the anterior and posterior sides of node cells, with anterior baMTs extending horizontally and posterior baMTs extending vertically. This asymmetry became evident after polarization of the PCP core protein Vangl1 and before the posterior positioning of the basal body, and it also required the PCP core proteins Prickle and dachsous. Our results suggest that the asymmetry in baMT organization may play a role in correct positioning of the basal body for left-right symmetry breaking.

Gse1, a component of the CoREST complex, is required for placenta development in the mouse.

Gse1 is a component of the CoREST complex that acts as an H3K4 and H3K9 demethylase and regulates gene expression. Here, we examined the expression and role of Gse1 in mouse development. Gse1 is expressed in male and female germ cells and plays both maternal and zygotic roles. Thus, maternal deletion of Gse1 results in a high incidence of prenatal death, and zygotic deletion leads to embryonic lethality from embryonic day 12.5 (E12.5) and perinatal death. Gse1 is expressed in the junctional zone and the labyrinth of the developing placenta. Gse1 mutant (Gse1Δex3/Δex3) placenta begins to exhibit histological defects from E14.5, being deficient in MCT4+ syncytiotrophoblast II. The number of various cell types was largely maintained in the mutant placenta at E10.5, but several genes were upregulated in giant trophoblasts at E10.5. Placenta-specific deletion of Gse1 with Tat-Cre suggested that defects in Gse1Δex3/Δex3 embryos are due to placental function deficiency. These results suggest that Gse1 is required for placental development in mice, and in turn, is essential for embryonic development.

Maternal epigenetic factors in embryonic and postnatal development.

Maternal factors present in oocytes and surrounding granulosa cells influence early development of embryos. In this study, we searched for epigenetic regulators that are expressed in oocytes and/or granulosa cells. Some of the 120 epigenetic regulators examined were expressed specifically in oocytes and/or granulosa cells. When their expression was examined in young versus aged oocytes or granulosa cells, many were significantly up- or downregulated in aged cells. The maternal role of six genes in development was investigated by generating oocyte-specific knock-out (MKO) mice. Two genes (Mllt10, Kdm2b) did not show maternal effects on later development, whereas maternal effects were evident for Kdm6a, Kdm4a, Prdm3, and Prdm16 for MKO female mice. Offspring from Kdm6a MKO mice underwent perinatal lethality at a higher rate. Pups derived from Prdm3;Prdm16 double MKO showed a higher incidence of postnatal death. Finally, embryos derived from Kdm4a MKO mice showed early developmental defects as early as the peri-implantation stage. These results suggest that many of maternal epigenetic regulators undergo differential expression upon aging. Some, such as Kdm4a, Kdm6a, Prdm3, and Prdm16, have maternal role in later embryonic or postnatal development.

CFAP53 regulates mammalian cilia-type motility patterns through differential localization and recruitment of axonemal dynein components.

Motile cilia can beat with distinct patterns, but how motility variations are regulated remain obscure. Here, we have studied the role of the coiled-coil protein CFAP53 in the motility of different cilia-types in the mouse. While node (9+0) cilia of Cfap53 mutants were immotile, tracheal and ependymal (9+2) cilia retained motility, albeit with an altered beat pattern. In node cilia, CFAP53 mainly localized at the base (centriolar satellites), whereas it was also present along the entire axoneme in tracheal cilia. CFAP53 associated tightly with microtubules and interacted with axonemal dyneins and TTC25, a dynein docking complex component. TTC25 and outer dynein arms (ODAs) were lost from node cilia, but were largely maintained in tracheal cilia of Cfap53-/- mice. Thus, CFAP53 at the base of node cilia facilitates axonemal transport of TTC25 and dyneins, while axonemal CFAP53 in 9+2 cilia stabilizes dynein binding to microtubules. Our study establishes how differential localization and function of CFAP53 contributes to the unique motion patterns of two important mammalian cilia-types.

Increased Chymase-Positive Mast Cells in High-Grade Mucoepidermoid Carcinoma of the Parotid Gland.

It has long been known that high-grade mucoepidermoid carcinoma (MEC) has a poor prognosis, but the detailed molecular and biological mechanisms underlying this are not fully understood. In the present study, the pattern of chymase-positive mast cells, as well as chymase gene expression, in high-grade MEC was compared to that of low-grade and intermediate-grade MEC by using 44 resected tumor samples of MEC of the parotid gland. Chymase expression, as well as chymase-positive mast cells, was found to be markedly increased in high-grade MEC. Significant increases in PCNA-positive cells and VEGF gene expression, as well as lymphangiogenesis, were also confirmed in high-grade MEC. Chymase substrates, such as the latent transforming growth factor-beta (TGF-ß) 1 and pro-matrix metalloproteinase (MMP)-9, were also detected immunohistologically in high-grade MEC. These findings suggested that the increased chymase activity may increase proliferative activity, as well as metastasis in the malignant condition, and the inhibition of chymase may be a strategy to improve the poor prognosis of high-grade MEC of the parotid gland.

Visceral adipose tissue quality was associated with nonalcoholic fatty liver disease, independent of its quantity.

BACKGROUND AND AIMS: Nonalcoholic fatty liver disease (NAFLD) is a serious liver disease. Recent studies have shown that both visceral adipose tissue (VAT) quantity and density (as an indirect measure of quality) are associated with metabolic profiles. Therefore, we investigated the association between VAT quantity and quality, and the prevalence and incidence of NAFLD. METHODS AND RESULTS: In this cross-sectional, retrospective cohort study, the prevalence and incidence of NAFLD were analyzed in 627 and 360 middle-aged subjects, respectively. VAT was evaluated using an unenhanced computed tomography scan, while NAFLD was evaluated using ultrasonography. The VAT area was normalized to the square value of the subjects' height in meters, the visceral fat area (VFA) index. The VAT density was described as the visceral fat density (VFD). The VFA index and VFD had an interaction effect on the prevalence of NAFLD (P = 0.0059). The VFA index (adjusted odds ratio [OR], 1.04; 95% confidence interval [CI], 1.01-1.07; P = 0.0145, per 1.0 cm2/m2) and the VFD (OR, 0.90; 95% CI, 0.84-0.96; P = 0.0026, per 1.0 Hounsfield unit [HU]) were independently associated with the prevalence of NAFLD. In our cohort, 36 subjects developed NAFLD. The VFD (adjusted hazards ratio [HR], 0.84; 95% CI, 0.77-0.91; P < 0.0001, per 1.0 HU) was independently associated with the incidence of NAFLD, whereas the VFA index was not. CONCLUSION: Both the VFA index and VFD were independently associated with NAFLD prevalence. The VFD might be more related to the incidence of NAFLD than the VFA index.

Effects of Longitudinal Changes in Lifestyle-Related Risk Factors on the Incidence of Major Adverse Cardiac and Cerebrovascular Disease in Young Adults.

Metabolic syndrome (MetS) is one focus of healthcare system reform in Japan. We examined the effects of changes in individual risk factors over time on the incidence of major adverse cardio-cerebrovascular events (MACCE) in adults under the age of 50 years. Study participants under the age of 50 with neither hypertension nor hyperglycemia at baseline were analyzed. We used a parametric proportional hazard model to determine the effect of changes in abdominal circumference, blood pressure, serum lipids, and blood glucose on the incidence of MACCE.A total of 6,125 women and 6,403 men were subject to the analyses. The incidence rate of MACCE per 1,000 person-years was 1.17 for women and 2.42 for men. In men under the age of 50, an increase in abdominal circumference was associated with an increase in MACCE incidence (hazard ratio per 1 cm increase: 1.10; 95% confidence interval [CI], 1.04-1.17), whereas no statistically significant association was observed in women. Compared with Visit 1, if the abdominal circumference increased by 4 cm at Visit 3, the hazard ratio for developing MACCE was approximately 1.5 (hazard ratio 1.48; 95% CI, 1.18-1.86). In men under the age of 50, increases in abdominal circumference and systolic blood pressure were associated with an increased risk of developing MACCE, regardless of the degree of obesity at baseline. Therefore, encouraging young adults to improve their health before developing MetS may reduce the risk of MACCE.

Relationship Between Body Mass Index and Incidence of Atrial Fibrillation in Young Japanese Men　- The Nishimura Health Survey.

BACKGROUND: The associations between body mass index (BMI) and incidence of atrial fibrillation (AF) in young men are scarce, especially in Asian countries, given the differences in BMI between Asians and Western populations.Methods and Results:This study analyzed 17,865 middle-aged Japanese men without AF from a cohort of employees undergoing annual health examinations. AF incidence was evaluated during a follow-up period (median 4.0 years, interquartile range 2.0-7.1 years). Among young men aged 30-49 years, AF incidence was 0.64/1,000 person-years, whereas it was 2.54/1,000 and 7.60/1,000 person-years among men aged 50-59 and ≥60 years, respectively. Multivariable Cox regression analysis among young men revealed age (hazard ratio [HR] 3.28 by 10-years' increase, 95% confidence interval [CI] 1.72-6.25, P<0.001), BMI (BMI-quadratic, HR 1.01, 95% CI 1.00-1.01, P<0.001, BMI-linear, HR 0.95, 95% CI 0.86-1.05, P=0.33), and electrocardiogram (ECG) abnormalities, such as PQ prolongation, supraventricular beat, and p wave abnormality (HR 8.79, 95% CI 3.05-25.32, P<0.001), were significantly associated with AF incidence. There was a reverse J-shaped association between BMI and AF incidence in young men, whereby the presence of ECG abnormality inversely influenced the BMI-incident AF relationship. A linear association between BMI and AF incidence in men aged 50-59 and ≥60 years was present. CONCLUSIONS: AF incidence displays a reverse J-shaped relationship with BMI in young men, but a linear association in men aged ≥50 years. The paradoxical relationship seen in young men only may reflect atrial electrical or structural abnormalities.

Ciliogenesis-coupled accumulation of IFT-B proteins in a novel cytoplasmic compartment.

Cluap1/IFT38 is a ciliary protein that belongs to the IFT-B complex and is required for ciliogenesis. In this study, we have examined the behaviors of Cluap1 protein in nonciliated and ciliated cells. In proliferating cells, Cluap1 is located at the distal appendage of the mother centriole. When cells are induced to form cilia, Cluap1 is found in a novel noncentriolar compartment, the cytoplasmic IFT spot, which mainly exists once in a cell. Other IFT-B proteins such as IFT46 and IFT88 are colocalized in this spot. The cytoplasmic IFT spot is present in mouse embryonic fibroblasts (MEFs) but is absent in ciliogenesis-defective MEFs lacking Cluap1, Kif3a or Odf2. The cytoplasmic IFT spot is also found in mouse embryos but is absent in the Cluap1 mutant embryo. When MEFs are induced to form cilia, the cytoplasmic IFT spot appears at an early step of ciliogenesis but starts to disappear when ciliogenesis is mostly completed. These results suggest that IFT-B proteins such as Cluap1 accumulate in a previously undescribed cytoplasmic compartment during ciliogenesis.

Relationship between nonalcoholic fatty liver disease and muscle quality as well as quantity evaluated by computed tomography.

BACKGROUND & AIMS: Sarcopenia is reported to be associated with nonalcoholic fatty liver disease (NAFLD). Evaluation of skeletal muscle attenuation and area by computed tomography (CT) may represent a promising approach for evaluation of the risk of NAFLD. We examined the association between skeletal muscle characteristics and NAFLD and investigated the combined effect of these parameters on the prevalence of NAFLD. METHODS: In this cross-sectional study, we analysed data from 632 middle-aged Japanese subjects without daily alcohol intake (353 men and 279 women) from a cohort of employees undergoing annual health examinations. The cross-sectional skeletal muscle area was evaluated on the basis of CT data at the level of the third lumbar vertebrae, and the skeletal muscle index (SMI) and density (SMD) were calculated. The subjects were divided into four study groups according to their SMI and SMD relative to median values. RESULTS: One hundred forty men and forty-three women had NAFLD. Total SMI (odds ratio [OR] per 1.0 cm2 /kg/m2 increase 0.43, 95% confidence interval [CI] 0.29-0.64 in men and OR 0.21, 95% CI 0.10-0.42 in women) and total SMD (OR, per 1.0 Hounsfield Unit increase 0.88, 95% CI 0.83-0.93 in men and 0.88, 0.82-0.95 in women) were significantly associated with the prevalence of NAFLD after adjusting for covariates. The subgroup with simultaneous presence of low SMI and low SMD was associated with a significantly higher prevalence of NAFLD compared with other groups. CONCLUSIONS: Both SMI and SMD are independently associated with the prevalence of NAFLD.

Trunk muscle quality and quantity predict the development of metabolic syndrome and the increase in the number of its components in individuals without metabolic syndrome.

BACKGROUND AND AIMS: The metabolic syndrome has been reported by cross-sectional studies to have an association with skeletal muscle quality and quantity. Using a longitudinal study design, this study aimed to explicate the association between muscle characteristics assessed with computed tomography (CT) and the incidence and progression of metabolic syndrome. METHODS AND RESULTS: In this retrospective study on a cohort of employees undergoing annual physical examinations, we evaluated data from 554 participants without metabolic syndrome. The cross-sectional skeletal muscle area was determined based on CT data at the level of the third lumbar vertebra, and the skeletal muscle density (SMD) and skeletal muscle index (SMI) were measured. The participants were divided into four study groups according to the sex-specific median values for SMI and SMD. We followed the participants for a mean period of 3.1 years. In the sex- and age-adjusted model, SMI and SMD had an interaction effect on the longitudinal change in number of metabolic syndrome components (ß = -0.074, p = 0.0727). Multiple regression analyses revealed that both low SMI and SMD was significantly associated with the change (ß = 0.131, p = 0.0281), whereas the low SMI and high SMD, and high SMI and low SMD were not. Both low SMI and SMD (hazard ratio (HR), 2.42; 95% confidence interval, 1.28-4.78) showed an increased adjusted HR for incident metabolic syndrome. CONCLUSION: The participants with both low quality and quantity of skeletal muscles were associated with the incidence and progression of metabolic syndrome, whereas those with only low quantity or quality of skeletal muscles were not.

Cluap1 localizes preferentially to the base and tip of cilia and is required for ciliogenesis in the mouse embryo.

Qilin is one of several genes in zebrafish whose mutation results in cystic kidney. We have now studied the role of its mouse ortholog, Cluap1, in embryonic development by generating Cluap1 knockout (Cluap1-/-) mice. Cluap1-/- embryos died mid-gestation manifesting impairment of ciliogenesis in various regions including the node and neural tube. The basal body was found to be properly docked to the apical membrane of cells in the mutant, but the axoneme failed to grow. Cluap1 is a ciliary protein and is preferentially localized at the base and tip of cilia. Hedgehog signaling, as revealed with a Pacthed1-lacZ reporter gene, was lost in Cluap1-/- embryos at embryonic day (E) 8.5 but was ectopically expanded at E9.0. The Cluap1 knockout embryos also failed to manifest left-right asymmetric expression of Nodal in the lateral plate, most likely as a result of the loss of Hedgehog signaling in node crown cells that in turn leads to pronounced down-regulation of Gdf1 expression in these cells. Crown cell-specific restoration of Cluap1 expression rescued Gdf1 expression in crown cells and left-sided Nodal expression in the lateral plate of mutant embryos. Our results suggest that Cluap1 contributes to ciliogenesis by regulating the intraflagellar transport (IFT) cycle at the base and tip of the cilium.

Proposal for a novel classification of benign parotid tumors based on localization.

OBJECTIVE: Postoperative facial nerve paralysis is the most problematic complication after surgical treatment of parotid tumors. Localization of tumors is highly relevant for the surgical approach, but existing classification systems do not focus on the association between localization and surgical technique. Therefore, we created a new localization-based classification system for benign parotid tumors and investigated the characteristics of tumors in each localization and the frequency of postoperative facial nerve paralysis by retrospectively applying the classification to previous cases. METHODS: First, we defined 6 portions of the parotid gland (upper, U; lower, L; posterior, P; anterior, A; superficial, S; deep, D) by dividing the transverse plane into an upper and lower portion at the mandibular marginal branch, the longitudinal plane into a posterior and anterior portion at the midline of the parotid anteroposterior diameter, and the sagittal plane into a superficial and deep portion along the course of the facial nerve. Then, we defined 8 locations by combining the 6 portions in all possible ways (i.e., U-P-S, U-P-D, U-A-S, U-A-D, L-P-S, L-P-D, L-A-S, L-A-D). We used this classification to define the tumor localization in 948 patients who had undergone partial superficial parotidectomy for benign parotid tumors and then investigated the incidence, histopathological type, signs/symptoms, diagnosis, surgery, and complications in each area. RESULTS: Pleomorphic adenomas comprised approximately 70% of tumors in the upper portion but only approximately 35% in the lower portion. The rate of postoperative facial nerve paralysis was significantly higher for tumors in deep locations than in superficial locations (33.9% vs 14.9%, respectively), and the odds ratios for postoperative facial nerve paralysis in the U-P-D and U-A-D locations were 7.6 and 4.8 compared to the L-P-S location. When maximum diameter, operation time, bleeding volume, sex (reference: female), and age were added as control variables, the odds ratios were 4.2 and 3.0. CONCLUSION: Determining tumor localization preoperatively with the new localization-based classification of parotid tumors is helpful not only for predicting the histopathological type but also for predicting surgical complications, particularly postoperative facial nerve paralysis.

Trunk muscle quality and quantity are associated with renal volume in nondiabetic people.

Background: Renal disease is a major problem in terms of community health and the economy. Skeletal muscle is involved in crosstalk with the kidney. We therefore investigated the relationship between muscle quality and quantity, and renal parenchymal volume (RPV). Methods: The association between the parameters of skeletal muscle and RPV/body surface area (BSA) was analyzed by computed tomography in 728 middle-aged participants without kidney disease or diabetes mellitus in a cross-sectional study. A retrospective cohort study of 68 participants was undertaken to analyze the association between changes in RPV/BSA and muscle parameters. Parameter change was calculated as follows: parameter at the follow-up examination/parameter at the baseline examination. The normal attenuation muscle (NAM) and low attenuation muscle (LAM) were identified by Hounsfield Unit thresholds of +30 to +150, and -29 to +29, respectively. Results: Positive correlations were found between estimated glomerular filtration rate and RPV/BSA (r = 0.451, P < .0001). Multiple regression analyses revealed that the NAM index was positively related to RPV/BSA (ß = 0.458, P < .0001), whereas the LAM index was negatively related to RPV/BSA (ß = -0.237, P < .0001). In this cohort study, a change in the LAM index was independently associated with a change in RPV/BSA (ß = -0.349, P = .0032). Conclusion: Both trunk muscle quantity and quality were associated with renal volume related to renal function in nondiabetic people. An increase in low quality muscle volume might be related to a decrease in renal volume.

Immotile cilia mechanically sense the direction of fluid flow for left-right determination.

Immotile cilia at the ventral node of mouse embryos are required for sensing leftward fluid flow that breaks left-right symmetry of the body. However, the flow-sensing mechanism has long remained elusive. In this work, we show that immotile cilia at the node undergo asymmetric deformation along the dorsoventral axis in response to the flow. Application of mechanical stimuli to immotile cilia by optical tweezers induced calcium ion transients and degradation of Dand5 messenger RNA (mRNA) in the targeted cells. The Pkd2 channel protein was preferentially localized to the dorsal side of immotile cilia, and calcium ion transients were preferentially induced by mechanical stimuli directed toward the ventral side. Our results uncover the biophysical mechanism by which immotile cilia at the node sense the direction of fluid flow.

[How much pressure is applied on the eye balls during craniotomy?: Measurement with FlexiForce contact surface force sensor].

BACKGROUND: Visual loss following craniotomy is a serious postoperative complication in which elevation of ocular pressure during retraction of the skin flap may cause retinal ischemia. We reported that continuous monitoring of extraocular pressure with the FlexiForce sensor may avoid excessive skin flap retraction during craniotomy and thus prevent ocular complications. METHODS: Between January 2008 and December 2011, we analyzed data from 46 consecutive patients for whom continuous monitoring of extraocular pressure with FlexiForce sensor was performed. This sensor continuously displays the compressive force, allowing surgeons to check values on the monitor at any time. An alarm sounds if 50 gf is exceeded. We analyzed the temporal course of extraocular pressure and the relationship with patient characteristics. RESULT: No visual complications were encountered in this patient series. Maximum compressive force during craniotomy was 35.8±27.2 gf, with increases typically seen when surgeons used hooks or drills. However, due to the alarm, no prolonged periods of high force were noted in any patient. Effective methods for reducing force were: (1) taking off hooks on the compressive side; (2) changing the direction of hook tension; and (3) placing cushions such as gauze under the side of the skin flap. Maximum compressive force during microsurgery was 21.8±18.4 gf, and correlated with the beginning force of microsurgery. CONCLUSION: Compressive force was greatly reduced compared to the force reported previously. The etiologies of visual disability are not fully understood, but this sensor may be helpful in reducing extraocular compression.

Management and outcome of parotid mucoepidermoid carcinoma by histological grade: A 21-year review.

Objective: Mucoepidermoid carcinoma (MEC) is the most common malignancy of the parotid gland, but the outcome depends on the histological grade. Therefore, the aim of this study was to evaluate MEC on the basis of histological grade. Study Design: Retrospective analysis. Methods: We performed a retrospective analysis of data from patients whose initial treatment for MEC of the parotid gland was performed at our department between 1999 and 2021. We examined the association between the Armed Forces Institute of Pathology (AFIP) grade and outcome. Results: The AFIP grades were as follows: low, 26 cases; intermediate, 9 cases; and high, 31 cases. About 50% of cases were correctly diagnosed as malignant, and both grade and histology were accurately determined by fine-needle aspiration cytology in 20% of cases. The 5-year disease-free survival rate was 95.5% and 53.8% in the low-/intermediate- and high-grade cases, respectively. In the high-grade group, cases with recurrence were found to have a higher rate of lymph nodes metastasis than cases without recurrence. Furthermore, in this high-grade group, total sacrifice of the facial nerve did not reduce local recurrence. However, radical resection in the cases without tumor invasion to the nerve has decreased the local recurrence rate. The CRTC1-MAML2 fusion gene was expressed in 42.3% of low-/intermediate- and 14.3% of high-grade cases. Conclusions: The survival rate in MEC was quite different between the low-/intermediate- and high-grade cases. However, the rate of correct assessment of the grade by fine-needle aspiration cytology was poor. In high-grade cases, total sacrifice of the facial nerve may improve the rate of local recurrence in cases without invasion of the main trunk of the nerve. Expression of the CRTC1-MAML2 fusion gene could be helpful in not only the assessment of grade but the prediction of recurrence. Level of Evidence: 4.

Management for Warthin Tumor of the Parotid Gland: Surgery or Observation. A 21-Year Retrospective Study of 387 Cases.

PURPOSE: This study investigated the characteristics, diagnosis, and treatment of Warthin tumors (WTs) to explore the possibility of managing patients by observation. METHODS: We reviewed the records of 1167 patients with benign parotid tumors who were seen in our department between September 1999 and April 2021. Among them, 387 cases were WT and 668 cases were pleomorphic adenoma. We evaluated preoperative diagnoses of WT by symptoms/signs, fine-needle aspiration cytology (FNAC), imaging, such as ultrasonography and magnetic resonance imaging, and technetium-99m pertechnetate (Tc-99m) scintigraphy. Fisher's exact test and the Mann-Whitney U test were used in statistical analyses. RESULTS: Warthin tumors were treated by surgery in 238 cases and follow-up in 149 cases. The 238 patients were diagnosed as WT at the final pathology after surgery. Among them, 172 patients (72.3%) were determined as benign histological type by preoperative FNAC; in these 172 patients, 170 (71.4%) were correctly diagnosed as WT in the final pathology. Preoperative Tc-99m scintigraphy was performed in 69 patients diagnosed with WT by final pathology or FNAC, and the positive rate of Tc-99m scintigraphy in WT was 75.4%. CONCLUSIONS: Combining FNAC and Tc-99m scintigraphy, as well as considering clinical findings, enables the diagnosis of WT in most cases. In particular, WT is more common in the elderly, grows more slowly, and is less likely to be malignant. Therefore, if WT can be diagnosed preoperatively with a high rate of correct diagnosis, it could be an accurate and effective means of managing patients through follow-up without surgery.

Lnk deletion reinforces the function of bone marrow progenitors in promoting neovascularization and astrogliosis following spinal cord injury.

Lnk is an intracellular adaptor protein reported as a negative regulator of proliferation in c-Kit positive, Sca-1 positive, lineage marker-negative (KSL) bone marrow cells. The KSL fraction in mouse bone marrow is believed to represent a population of hematopoietic and endothelial progenitor cells (EPCs). We report here that, in vitro, Lnk(-/-) KSL cells form more EPC colonies than Lnk(+/+) KSL cells and show higher expression levels of endothelial marker genes, including CD105, CD144, Tie-1, and Tie2, than their wild-type counterparts. In vivo, the administration of Lnk(+/+) KSL cells to a mouse spinal cord injury model promoted angiogenesis, astrogliosis, axon growth, and functional recovery following injury, with Lnk(-/-) KSL being significantly more effective in inducing and promoting these regenerative events. At day 3 following injury, large vessels could be observed in spinal cords treated with KSL cells, and reactive astrocytes were found to have migrated along these large vessels. We could further show that the enhancement of astrogliosis appears to be caused in conjunction with the acceleration of angiogenesis. These findings suggest that Lnk deletion reinforces the commitment of KSL cells to EPCs, promoting subsequent repair of injured spinal cord through the acceleration of angiogenesis and astrogliosis.

Concurrent vasculogenesis and neurogenesis from adult neural stem cells.

RATIONALE: Recent reports have demonstrated that signals from vascular endothelial cells are necessary for organogenesis that may precede vasculogenesis. However, the origin of these neovascular cells in regenerating tissue has not been clarified. OBJECTIVE: Here we tested the hypothesis that adult neural stem cells (NSCs) can differentiate into vascular lineage, as well as neural lineage, in the process of collaborative organogenesis. METHODS AND RESULTS: NSCs, clonally isolated from mouse brain, were shown to develop endothelial and smooth muscle phenotypes in vitro. To elucidate whether NSCs can simultaneously differentiate into vascular and neural cells in vivo, genetically labeled NSCs were administered to mice with unilateral sciatic nerve crush injury or operatively induced brain and myocardial ischemia. Two weeks later, necropsy examination disclosed recruitment of the labeled NSCs to sites of injury differentiating into vascular cells (endothelial cells and vascular smooth muscle cells) and Schwann cells in regenerating nerve. Similarly, NSC-derived vascular cells/astrocytes and endothelial cells were identified in ischemic brain tissue and capillaries in myocardium 2 weeks following transplantation, respectively. CONCLUSIONS: These findings, concurrent vasculogenesis and neurogenesis from a common stem cell, suggest that certain somatic stem cells are capable of differentiating into not only somatic cells of identity but also into vascular cells for tissue regeneration.