Adaptive metabolic response to short-term intensive fasting.

BACKGROUND & AIMS: Short-term intensive fasting (STIF), known as beego in Chinese phonetic articulation, has been practiced for more than two thousand years. However, the potential risk of STIF and the body's response to the risk have not been adequately evaluated. This study aims to address this issue, focusing on the STIF-triggered metabolic response of the liver and kidney. METHODS: The STIF procedure in the clinical trial includes a 7-day water-only intensive fasting phase and a 7-day gradual refeeding phase followed by a regular diet. The intensive fasting in humans was assisted with psychological induction. To gain insights not available in the clinical trial, we designed a STIF program for mice that resulted in similar phenotypes seen in humans. Plasma metabolic profiling and examination of gene expression as well as liver and kidney function were performed by omics, molecular, biochemical and flow cytometric analyses. A human cell line model was also used for mechanistic study. RESULTS: Clinically significant metabolites of fat and protein were found to accumulate during the fasting phase, but they were relieved after gradual refeeding. Metabolomics profiling revealed a universal pattern in the consumption of metabolic intermediates, in which pyruvate and succinate are the two key metabolites during STIF. In the STIF mouse model, the accumulation of metabolites was mostly counteracted by the upregulation of catabolic enzymes in the liver, which was validated in a human cell model. Kidney filtration function was partially affected by STIF but could be recovered by refeeding. STIF also reduced oxidative and inflammatory levels in the liver and kidney. Moreover, STIF improved lipid metabolism in mice with fatty liver without causing accumulation of metabolites after STIF. CONCLUSIONS: The accumulation of metabolites induced by STIF can be relieved by spontaneous upregulation of catabolic enzymes, suggesting an adaptive and protective metabolic response to STIF stress in the mammalian body.

Short-term intensive fasting enhances the immune function of red blood cells in humans.

BACKGROUND: Fasting is known to influence the immune functions of leukocytes primarily by regulating their mobilization and redistribution between the bone marrow and the peripheral tissues or circulation, in particular via relocalization of leukocytes back in the bone marrow. However, how the immune system responds to the increased risk of invasion by infectious pathogens with fewer leukocytes in the peripheral blood during fasting intervention remains an open question. RESULTS: We used proteomic, biochemical and flow cytometric tools to evaluate the impact of short-term intensive fasting (STIF), known as beego, on red blood cells by profiling the cells from the STIF subjects before and after 6 days of fasting and 6 days of gradual refeeding. We found that STIF, by triggering the activation of the complement system via the complement receptor on the membrane of red blood cells, boosts fairly sustainable function of red blood cells in immune responses in close relation to various pathogens, including viruses, bacteria and parasites, particularly with the pronounced capacity to defend against SARS-CoV-2, without compromising their oxygen delivery capacity and viability. CONCLUSION: STIF fosters the immune function of red blood cells and therefore, it may be considered as a nonmedical intervention option for the stronger capacity of red blood cells to combat infectious diseases.

Photoalignment of sub-micrometer periodic liquid crystal polarization grating by using the optical imprinting method.

Photoalignment of liquid crystal polarization grating based on optical imprinting is a promising technique for polarization grating mass production. However, when the period of the optical imprinting grating is in the sub-micrometer level, the zero-order energy from the master grating will become high, and it will strongly affect the photoalignment quality. This paper proposes a double-twisted polarization grating structure to eliminate the zero-order disturbance of master grating and gives the design method. Based on the designed results, a master grating was prepared, and the optically imprinted photoalignment of polarization grating with a period of 0.5µm was fabricated. This method has the advantages of high efficiency and significantly greater environmental tolerance than the traditional polarization holographic photoalignment methods. It has the potential to be used for large-area polarization holographic gratings production.

Liquid-crystal-polymer binary diffractive optical elements with a sub-micrometer feature size realized by a contact polarization holography.

In this Letter, a contact polarization holographic photoalignment method is proposed. In the holographic recording, a phase mask is contacted with a photoalignment film, making light carrying wavefront information interfere with reference light in the near-field region to realize polarization holographic pattern recording with a sub-micrometer feature size. The relevant theoretical derivation is given, and holographic recording of a 0.4 µm feature-size phase mask is realized. The proposed method can conveniently realize liquid-crystal binary diffractive optical elements with a sub-micrometer feature size. Off-axis diffraction can also be realized by superimposing the grating information by changing the angle between the substrate and the interference light.

Autocrine pro-legumain promotes breast cancer metastasis via binding to integrin αvß3.

Tumor metastasis is the leading cause of cancer-associated mortality. Unfortunately, the underlying mechanism of metastasis is poorly understood. Expression of legumain (LGMN), an endo-lysosomal cysteine protease, positively correlates with breast cancer metastatic progression and poor prognosis. Here, we report that LGMN is secreted in the zymogen form by motile breast cancer cells. Through binding to cell surface integrin αvß3 via an RGD motif, the autocrine pro-LGMN activates FAK-Src-RhoA signaling in cancer cells and promotes cancer cell migration and invasion independent of LGMN protease activity. Either silencing LGMN expression or mutationally abolishing pro-LGMN‒αvß3 interaction significantly inhibits cancer cell migration and invasion in vitro and breast cancer metastasis in vivo. Finally, we developed a monoclonal antibody against LGMN RGD motif, which blocks pro-LGMN‒αvß3 binding, and effectively suppresses cancer cell migration and invasion in vitro and breast cancer metastasis in vivo. Thus, disruption of pro-LGMN‒integrin αvß3 interaction may be a potentially promising strategy for treating breast cancer metastasis.

Controlled Release of TGF-ß3 for Effective Local Endogenous Repair in IDD Using Rat Model.

Introduction: Intervertebral disc (IVD) degeneration (IDD) is one of the most widespread musculoskeletal diseases worldwide and remains an intractable clinical challenge. Currently, regenerative strategies based on biomaterials and biological factors to facilitate IVD repair have been widely explored. However, the harsh microenvironment, such as increased ROS and acidity, of the degenerative region impedes the efficiency of IVD repair. Here, an intelligent biodegradable nanoplatform using hollow manganese dioxide (H-MnO2) was developed to modulate the degenerative microenvironment and release transforming growth factor beta-3 (TGF-ß3), which may achieve good long-term therapeutic effects on needle puncture-induced IDD. Methods: Surface morphology and elemental analysis of the MnO2 nanoparticles (NPs) were performed by transmission electron microscopy and an energy-dispersive X-ray spectroscopy detector system, respectively. The biological effects of MnO2 loaded with TGF-ß3 (TGF-ß3/MnO2) on nucleus pulposus cells (NPCs) were assessed via cytoskeleton staining, EdU staining, qPCR and immunofluorescence. The efficacy of TGF-ß3/MnO2 on needle puncture-induced IDD was further examined using MRI and histopathological and immunohistochemical staining. Results: The MnO2 NPs had a spherical morphology and hollow structure that dissociated in the setting of a low pH and H2O2 to release loaded TGF-ß3 molecules. In the oxidative stress environment, TGF-ß3/MnO2 was superior to TGF-ß3 and MnO2 NPs in the suppression of H2O2-induced matrix degradation, ROS, and apoptosis in NPCs. When injected into the IVDs of a rat IDD model, TGF-ß3/MnO2 was able to prevent the degeneration and promote self-regeneration. Conclusion: Use of an MnO2 nanoplatform for biological factors release to regulate the IDD microenvironment and promote endogenous repair may be an effective approach for treating IDD.

Integrin αEß7+ T cells direct intestinal stem cell fate decisions via adhesion signaling.

Intestinal stem cell (ISC) differentiation is regulated precisely by a niche in the crypt, where lymphocytes may interact with stem and transient amplifying (TA) cells. However, whether and how lymphocyte-stem/TA cell contact affects ISC differentiation is largely unknown. Here, we uncover a novel role of T cell-stem/TA cell contact in ISC fate decisions. We show that intestinal lymphocyte depletion results in skewed ISC differentiation in mice, which can be rescued by T cell transfer. Mechanistically, integrin αEß7 expressed on T cells binds to E-cadherin on ISCs and TA cells, triggering E-cadherin endocytosis and the consequent Wnt and Notch signaling alterations. Blocking αEß7-E-cadherin adhesion suppresses Wnt signaling and promotes Notch signaling in ISCs and TA cells, leading to defective ISC differentiation. Thus, αEß7+ T cells regulate ISC differentiation at single-cell level through cell-cell contact-mediated αEß7-E-cadherin adhesion signaling, highlighting a critical role of the T cell-stem/TA cell contact in maintaining intestinal homeostasis.

Mucin-Like Domain of Mucosal Addressin Cell Adhesion Molecule-1 Facilitates Integrin α4ß7-Mediated Cell Adhesion Through Electrostatic Repulsion.

The homing of lymphocytes from blood to gut-associated lymphoid tissue is regulated by interaction between integrin α4ß7 with mucosal vascular addressin cell adhesion molecule 1 (MAdCAM-1) expressed on the endothelium of high endothelial venules (HEVs). However, the molecular basis of mucin-like domain, a specific structure of MAdCAM-1 regulating integrin α4ß7-mediated cell adhesion remains obscure. In this study, we used heparan sulfate (HS), which is a highly acidic linear polysaccharide with a highly variable structure, to mimic the negative charges of the extracellular microenvironment and detected the adhesive behaviors of integrin α4ß7 expressing 293T cells to immobilized MAdCAM-1 in vitro. The results showed that HS on the surface significantly promoted integrin α4ß7-mediated cell adhesion, decreased the percentage of cells firmly bound and increased the rolling velocities at high wall shear stresses, which was dependent on the mucin-like domain of MAdCAM-1. Moreover, breaking the negative charges of the extracellular microenvironment of CHO-K1 cells expressing MAdCAM-1 with sialidase inhibited cell adhesion and rolling velocity of 293T cells. Mechanistically, electrostatic repulsion between mucin-like domain and negative charges of the extracellular microenvironment led to a more upright conformation of MAdCAM-1, which facilitates integrin α4ß7-mediated cell adhesion. Our findings elucidated the important role of the mucin-like domain in regulating integrin α4ß7-mediated cell adhesion, which could be applied to modulate lymphocyte homing to lymphoid tissues or inflammatory sites.

Curettage through a wide cortical window for treatment of a primary aneurysmal bone cyst of the patella.

A 27-year-old man presented with intermittent right knee pain for 1 year with no previous trauma. Physical examination revealed only tenderness over the patella. Typical fluid-fluid levels were visible on magnetic resonance imaging (MRI), which highly suggested aneurysmal bone cyst (ABC) of the patella. After removal of a large window of thin cortical bone, curettage and bone grafting followed by cerclage wiring was performed. Histology confirmed the initial diagnosis of primary ABC of the patella. At the final follow-up visit at 71 months after surgery, the patient had normal joint activity with no pain or evidence of recurrence. Previous publications indicated patellectomy in the initial series, but curettage and bone grafting have more recently provided excellent results and good graft incorporation in most cases, even for aggressive lesions. In our patient, thorough curettage and bone grafting through a wide cortical window followed by cerclage wiring fixation and figure-eight sutures was a successful treatment option for primary ABC of the patella without articular disruption.

A mutation that blocks integrin α4ß7 activation prevents adaptive immune-mediated colitis without increasing susceptibility to innate colitis.

BACKGROUND: ß7 integrins are responsible for the efficient recruitment of lymphocytes from the blood and their retention in gut-associated lymphoid tissues. Integrin α4ß7 binds MAdCAM-1, mediating rolling adhesion of lymphocytes on blood vessel walls when inactive and firm adhesion when activated, thereby controlling two critical steps of lymphocyte homing to the gut. By contrast, integrin αEß7 mediates the adhesion of lymphocytes to gut epithelial cells by interacting with E-cadherin. Integrin ß7 blocking antibodies have shown efficacy in clinical management of inflammatory bowel disease (IBD); however, fully blocking ß7 function leads to the depletion of colonic regulatory T (Treg) cells and exacerbates dextran sulfate sodium (DSS)-induced colitis by evoking aberrant innate immunity, implying its potential adverse effect for IBD management. Thus, a better therapeutic strategy targeting integrin ß7 is required to avoid this adverse effect. RESULTS: Herein, we inhibited integrin α4ß7 activation in vivo by creating mice that carry in their integrin ß7 gene a mutation (F185A) which from structural studies is known to lock α4ß7 in its resting state. Lymphocytes from ß7-F185A knock-in (KI) mice expressed α4ß7 integrins that could not be activated by chemokines and showed significantly impaired homing to the gut. The ß7-F185A mutation did not inhibit αEß7 activation, but led to the depletion of αEß7+ lymphocytes in the spleen and a significantly reduced population of αEß7+ lymphocytes in the gut of KI mice. ß7-F185A KI mice were resistant to T cell transfer-induced chronic colitis, but did not show an increased susceptibility to DSS-induced innate colitis, the adverse effect of fully blocking ß7 function. CONCLUSIONS: Our findings demonstrate that specific inhibition of integrin α4ß7 activation is a potentially better strategy than fully blocking α4ß7 function for IBD treatment.

Fever Promotes T Lymphocyte Trafficking via a Thermal Sensory Pathway Involving Heat Shock Protein 90 and α4 Integrins.

Fever is an evolutionarily conserved response that confers survival benefits during infection. However, the underlying mechanism remains obscure. Here, we report that fever promoted T lymphocyte trafficking through heat shock protein 90 (Hsp90)-induced α4 integrin activation and signaling in T cells. By inducing selective binding of Hsp90 to α4 integrins, but not ß2 integrins, fever increased α4-integrin-mediated T cell adhesion and transmigration. Mechanistically, Hsp90 bound to the α4 tail and activated α4 integrins via inside-out signaling. Moreover, the N and C termini of one Hsp90 molecule simultaneously bound to two α4 tails, leading to dimerization and clustering of α4 integrins on the cell membrane and subsequent activation of the FAK-RhoA pathway. Abolishment of Hsp90-α4 interaction inhibited fever-induced T cell trafficking to draining lymph nodes and impaired the clearance of bacterial infection. Our findings identify the Hsp90-α4-integrin axis as a thermal sensory pathway that promotes T lymphocyte trafficking and enhances immune surveillance during infection.

Static Compression Induces ECM Remodeling and Integrin α2ß1 Expression and Signaling in a Rat Tail Caudal Intervertebral Disc Degeneration Model.

STUDY DESIGN: A three-level rat tail caudal intervertebral disc (IVD) degeneration (IVDD) model was established to study effects of static compression on extracellular matrix (ECM) remodeling and integrin signaling in IVDs during IVDD. OBJECTIVE: The aim of this study was to investigate the effect of compression force on ECM remodeling and integrin signaling in IVDs during IVDD. SUMMARY OF BACKGROUND DATA: Integrins sense mechanical environment alteration via binding to ECM ligands and trigger intracellular signaling for pathological ECM remodeling during IVDD. However, the role of compression force in ECM remodeling and integrin signaling during IVDD remains elusive. METHODS: Compared with the classical one-level rat tail IVDD model that exerts axial stress on the 8th to 9th caudal vertebral bodies, a three-level model was established by using an Ilizarov-type apparatus to exert stress on the 7th to 10th caudal vertebral bodies in rat tails for four weeks. To exclude side effects from surgical stab injury on manipulated discs, intact coccygeal (Co) disc Co8-9 was analyzed. RESULTS: In three-level IVDD model, significant degeneration of the Co8-9 disc was observed. Quantitative real-time polymerase chain reaction (qRT-PCR) showed elevated mRNA expression of collagen types I, III, and V; matrix metalloproteinases (MMPs) 2, 3, 9, 13, 14; and decreased mRNA expression of collagen type II in Co8-9 disc. Compression loading altered the expression of integrin α2ß1 (upregulated) and α10ß1 (downregulated) in NP cells, and activated integrin downstream signaling. By contrast, one-level model showed more severe disc degeneration and ECM remodeling. Integrin α1, α2, α11, and ß1 were upregulated, whereas α10 was downregulated. Similar activation of integrin signaling was observed. CONCLUSION: Static compression altered collagen and MMP expression, and promoted ß1 integrin expression and signaling in IVD. Compared with one-level rat tail IVDD model, three-level model showed milder effects on disc degeneration, ECM remodeling, and integrin expression, suggesting one-level model might involve other causes that induce IVDD via mechanisms independent of compression force. LEVEL OF EVIDENCE: N/A.

Kindlin-3 Is Essential for the Resting α4ß1 Integrin-mediated Firm Cell Adhesion under Shear Flow Conditions.

Integrin-mediated rolling and firm cell adhesion are two critical steps in leukocyte trafficking. Integrin α4ß1 mediates a mixture of rolling and firm cell adhesion on vascular cell adhesion molecule-1 (VCAM-1) when in its resting state but only supports firm cell adhesion upon activation. The transition from rolling to firm cell adhesion is controlled by integrin activation. Kindlin-3 has been shown to bind to integrin ß tails and trigger integrin activation via inside-out signaling. However, the role of kindlin-3 in regulating resting α4ß1-mediated cell adhesion is not well characterized. Herein we demonstrate that kindlin-3 was required for the resting α4ß1-mediated firm cell adhesion but not rolling adhesion. Knockdown of kindlin-3 significantly decreased the binding of kindlin-3 to ß1 and down-regulated the binding affinity of the resting α4ß1 to soluble VCAM-1. Notably, it converted the resting α4ß1-mediated firm cell adhesion to rolling adhesion on VCAM-1 substrates, increased cell rolling velocity, and impaired the stability of cell adhesion. By contrast, firm cell adhesion mediated by Mn(2+)-activated α4ß1 was barely affected by knockdown of kindlin-3. Structurally, lack of kindlin-3 led to a more bent conformation of the resting α4ß1. Thus, kindlin-3 plays an important role in maintaining a proper conformation of the resting α4ß1 to mediate both rolling and firm cell adhesion. Defective kindlin-3 binding to the resting α4ß1 leads to a transition from firm to rolling cell adhesion on VCAM-1, implying its potential role in regulating the transition between integrin-mediated rolling and firm cell adhesion.