[Pouchitis showed complete response to ustekinumab].

Pouchitis is the most common long-term complication following ileal pouch-anal anastomosis (IPAA) in patients with ulcerative colitis. Although several agents, including probiotics, steroids, and immunomodulators, have been used, the treatment of pouchitis remains challenging. Owing to the proven efficacy of biological therapy in inflammatory bowel disease, there is now growing evidence suggesting the potential benefits of biological therapy in refractory pouchitis. Here, we report the case of a 64-year-old woman with pouchitis due to ulcerative colitis who was successfully treated with ustekinumab (UST). The patient developed ulcerative pancolitis at the age of 35. Total colectomy and IPAA with J-pouch anastomosis were performed when the patient was 47 years old. Ileotomy closure was performed 6 months later. Postoperatively, the patient developed steroid-dependent pouchitis. Three years later, she developed steroid-induced diabetes. The patient has been taking 3mg of steroid for 20 years;therefore, her lifetime total steroid dose was 21g. The patient had over 20 episodes of bloody diarrhea a day. The last pouchoscopy in 20XX-9 revealed inflammatory stenosis with deep ulcerations of the afferent limb just before the ileoanal pouch junction. In July 20XX, when we took over her treatment, the policy of treatment was to withdraw her from steroids. Pouchoscopy revealed a widened but still tight afferent limb through which the scope could easily pass, and the ileoanal pouch still showed erosive ileitis without ulcers. Thiopurine administration and steroid tapering were initiated. Steroid tapering increased the erythrocyte sedimentation rate (ESR). As ESR increased, her arthritis exacerbated. Six months after the end of steroid administration, the patient consented to UST treatment. On April 20XX+1, the patient received her first 260-mg UST infusion. At this point, she experienced 14-15 episodes of muddy bloody stools. She had no abdominal pain;however, she experienced shoulder pain. Gradually, UST affected both pouchitis and arthritis. UST treatment was continued at 90mg subcutaneously every 12 weeks without abdominal pain recurrence. Eight months after the first UST infusion, nonsteroidal anti-inflammatory drugs were no longer necessary for shoulder pain. Follow-up pouchoscopy performed 14 months after UST optimization revealed a normal afferent limb without ulcerations in either segment. Pouchitis remission was maintained for over 2 years.

p24 family Tango(1) at the endoplasmic reticulum exit site to organize cargo exit.

The p24 family of proteins have been regarded as cargo receptors for endoplasmic reticulum (ER) to Golgi transport; however, their precise functions have yet to be revealed. In this issue, Pastor-Pareja and colleagues (https://doi.org/10.1083/jcb.202309045) show that the interaction of these proteins with Tango1 is critical for their localization at the ER exit site (ERES) and efficient transport of secretory proteins in Drosophila.

Evaluation of the enhancement of photosynthetic rate in a komatsuna (Brassica rapa L. var. perviridis) canopy with upward lighting using an optical simulation in a plant factory with artificial light.

In a plant factory with artificial light (PFAL), upward lighting is expected to prevent senescence and decrease in the photosynthetic capacity of the lower leaves in the canopy. Upward lighting may also increase the photosynthetic rate of a canopy by improving its photosynthetic photon flux density (PPFD) distribution. However, the net photosynthetic rate (Pn) of leaves is lower when the abaxial surface is irradiated than that when the adaxial surface is irradiated. The aim of this study was to estimate the PPFD in a PFAL and the Pn of plants using three-dimensional plant models and optical simulation. First, we measured the Pn of komatsuna (Brassica rapa L. var. perviridis) leaves under different conditions of the proportion (pad ) of PPFD on the adaxial surface to total PPFD on both surfaces and developed an equation for the light response curve of photosynthesis considering pad . When PPFD was low, except when it was 30 and 70 µmol m-2 s-1, Pn increased as pad increased, because the absorptance also increased with pad . Under high PPFD conditions, Pn was maximized at 67-83% of pad because the light would be distributed more efficiently for photosynthesis. Next, using optical simulation and the developed equation, we estimated the photosynthetic rate of a komatsuna canopy (CPn) under downward and upward lighting. The CPn increased by 1.08-1.13 times by combining downward and upward lighting due to the increase in the photosynthetic photon flux (PPF) of light incident on the canopy and the decrease in the spatial variation of PPFD on the leaves in the canopy. As the depreciation of lamps for upward lighting accounts for 7.5-9.0% of the production cost in a PFAL, even if the depreciation of lamps for upward lighting increased, enhancement of CPn by upward lighting would be cost-effective. We performed optical simulations under 220 conditions and evaluated them using CPn as an index. Moreover, we provided the proportion of PPF of upward lighting that improved CPn and discussed the reason for this improvement. The result shows that optical simulation is useful for evaluating the lighting design in a PFAL and analyzing the effects of the lighting design on the light environment and photosynthesis.

Structural Analysis of Intracellular Lipid Radicals by LC/MS/MS Using a BODIPY-Based Profluorescent Nitroxide Probe.

Free radical-mediated lipid peroxidation (LPO) induces the formation of numerous lipid radicals, which contribute to the development of several oxidative diseases. To understand the mechanism of LPO in biological systems and the significance of these radicals, identifying the structures of individual lipid radicals is imperative. In this study, we developed an analytical method based on liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS) and a profluorescent nitroxide probe, N-(1-oxyl-2,2,6-trimethyl-6-pentylpiperidin-4-yl)-3-(5,5-difluoro-1,3-dimethyl-3H,5H-5l4-dipyrrolo[1,2-c:2',1'-f][1,3,2]diazaborinin-7-yl)propanamide (BDP-Pen), for the detailed structural analysis of lipid radicals. The MS/MS spectra of BDP-Pen-lipid radical adducts showed product ions and thus allow the prediction of the lipid radical structures and individual detection of isomeric adducts. Using the developed technology, we separately detected the isomers of arachidonic acid (AA)-derived radicals generated in AA-treated HT1080 cells. This analytical system is a powerful tool for elucidating the mechanism of LPO in biological systems.

Slippage- and load-induced changes in the crystalline orientation of oligo(3-methoxythiophene) powder to develop a gold-tone luster.

The achievement of molecular orientation control by rubbing and pressing poly(3-alkylthiophene)s is a powerful technique to improve the performance of organic electronic devices. We report here that the rubbing and pressing of blackish-brown 3-methoxythiophene oligomer powders yield layer and tablet samples with gold tones, respectively. Specular reflectivity, colorimetric, and X-ray diffraction measurements reveal that this gold tone is caused by an increase in the ratio of edge-on lamellar crystallites to face-on ones, which is promoted by rubbing/pressing. In contrast to the 3-alkylthiophene polymer, which develops a dominant face-on lamellar structure, rubbing of the 3-methoxythiophene oligomer increases the relative amount of edge-on lamellar crystallites to face-on lamellar ones. Furthermore, gold tone development in the tablet samples is limited to the near-surface area, despite the fact that pressure is also applied to the tablet bulk. These specific chemical events are explained by considering the repulsive interactions between the 3-methoxythiophene backbone and the functional groups on the surface of the substrate employed during the rubbing/pressing processes. Despite the lower applied pressure, gold tone development by rubbing is accompanied by a higher reflective property than by pressing because of the formation of larger relative amounts and sizes of edge-on lamellar crystallites, which are responsible for the gold tone.

Cargo receptor Surf4 regulates endoplasmic reticulum export of proinsulin in pancreatic ß-cells.

Insulin is an essential peptide hormone that maintains blood glucose levels. Although the mechanisms underlying insulin exocytosis have been investigated, the mechanism of proinsulin export from the endoplasmic reticulum (ER) remains unclear. Here, we demonstrated that Surf4, a cargo receptor homolog, regulates the ER export of proinsulin via its recruitment to ER exit sites (ERES). Under high-glucose conditions, Surf4 expression was upregulated, and Surf4 proteins mainly localized to the ER at a steady state and accumulated in the ERES, along with proinsulin in rat insulinoma INS-1 cells. Surf4-knockdown resulted in proinsulin retention in the ER and decreased the levels of mature insulin in secretory granules, thereby significantly reducing insulin secretion. Surf4 forms an oligomer and can physically interact with proinsulin and Sec12, essential for COPII vesicle formation. Our findings suggest that Surf4 interacts with proinsulin and delivers it into COPII vesicles for ER export in co-operation with Sec12 and COPII.

Remimazolam anesthesia for cardiac surgery with cardiopulmonary bypass: a case report.

BACKGROUND: Remimazolam has less cardiovascular depressant effects than propofol in non-cardiac surgical patients. However, the efficacy and safety of remimazolam in cardiac surgery with cardiopulmonary bypass (CPB) have not been reported. We present a case of successful anesthetic management using remimazolam in cardiac surgery with CPB. CASE PRESENTATION: A 76-year-old female was scheduled for mitral valve repair, tricuspid annuloplasty, maze procedure, and left atrial appendage closure. We used remimazolam in induction (6.0 mg/kg/h) and maintenance (0.6-1.0 mg/kg/h) of general anesthesia, and the bispectral index value was maintained in the range of 36 to 48 including the period of CPB. Hemodynamics, mixed venous oxygen saturation, and bilateral regional cerebral oxygen saturation were maintained within acceptable ranges. There was no intraoperative awareness/recall or serious complications associated with remimazolam throughout the perioperative period. CONCLUSIONS: Remimazolam can be used the same as other existing anesthetics in cardiac surgery with CPB.

Mitotic ER exit site dynamics: insights into blockade of secretion from the ER during mitosis.

How ER exit sites disassemble during mitosis is not well understood. Transport ANd Golgi Organization 1 (TANGO1, also known as MIA3), a cargo receptor originally identified for collagens, acts as a hub for ER exit site disassembly under the control of Casein Kinase 1 (CK1)-mediated phosphorylation and Protein Phosphatase 1 (PP1)-mediated dephosphorylation. Impaired dephosphorylation during mitosis induces ER exit site disassembly.

Sequence Type Changes Associated with Decreasing Macrolide-Resistant Mycoplasma pneumoniae, Japan.

We compared sequence types (STs) of Mycoplasma pneumoniae isolates from Japan during 2002-2019. ST3 and ST14 dominated during 2002-2016, and ST7 and ST33 dominated during 2018-2019. These STs were associated with a decrease in macrolide-resistant strains after an epidemic of infection with M. pneumoniae during 2011-2012.

Mitotic ER Exit Site Disassembly and Reassembly Are Regulated by the Phosphorylation Status of TANGO1.

Golgi fragmentation and ER exit site disassembly are considered to be the leading causes of the mitotic block of secretion from the ER. Although the mechanisms of Golgi fragmentation have been extensively characterized, ER exit block early in mitosis is not well understood. We previously demonstrated that TANGO1 organizes ER exit sites by directly interacting with Sec16. In this study, we identified TANGO1 as a regulator of ER exit site disassembly during mitosis. TANGO1 phosphorylation was observed to be coordinately regulated by a kinase (CK1) and a phosphatase (PP1). CK1-mediated TANGO1 phosphorylation reduces binding to Sec16, leading to the disassembly of ER exit sites. CK1 constantly phosphorylates TANGO1, whereas PP1-mediated dephosphorylation of TANGO1 decreases during mitosis. Thus, the phosphorylation status of TANGO1, which is controlled by balanced activities of the kinase CK1 and the phosphatase PP1, regulates the organization of ER exit sites during the cell cycle.

Reaction targets of antioxidants in azo-initiator or lipid hydroperoxide induced lipid peroxidation.

Lipid peroxidation (LPO) is reported to be involved in the pathogenesis of several oxidative diseases, and several therapeutic approaches using antioxidants have been proposed. LPO is thought to progress via a complicated series of multistep reactions suggesting that the activity of each antioxidant may be different, and depends on the reacting molecules. Hence, in this study, we evaluated the inhibitory mechanisms of several antioxidants toward arachidonic acid (AA) peroxidation induced by the azo initiator 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) or a lipid hydroperoxide, hydroperoxyoctadecadienoic acid (HpODE)/hemin. Edaravone, ferrostatin-1, TEMPO and trolox effectively inhibited the production of malondialdehyde (MDA) and several oxidised AAs generated in the AAPH-induced LPO because of their scavenging ability toward lipid peroxyl radicals. In contrast, ebselen and ferrostatin-1 showed strong antioxidative activity in the HpODE/hemin-induced peroxidation. Under this condition, ebselen and ferrostatin-1 were thought to reduce HpODE and its derived alkoxyl radicals to the corresponding lipid alcohols. In conclusion, we found that each antioxidant had different antioxidative activities that prevented the progression of LPO. We expect that these findings will contribute to the design of novel therapeutic strategies using an appropriate antioxidant targeted to each step of the development of oxidative stress diseases.

LTK is an ER-resident receptor tyrosine kinase that regulates secretion.

The endoplasmic reticulum (ER) is a key regulator of cellular proteostasis because it controls folding, sorting, and degradation of secretory proteins. Much has been learned about how environmentally triggered signaling pathways regulate ER function, but only little is known about local signaling at the ER. The identification of ER-resident signaling molecules will help gain a deeper understanding of the regulation of ER function and thus of proteostasis. Here, we show that leukocyte tyrosine kinase (LTK) is an ER-resident receptor tyrosine kinase. Depletion of LTK as well as its pharmacologic inhibition reduces the number of ER exit sites and slows ER-to-Golgi transport. Furthermore, we show that LTK interacts with and phosphorylates Sec12. Expression of a phosphoablating mutant of Sec12 reduces the efficiency of ER export. Thus, LTK-to-Sec12 signaling represents the first example of an ER-resident signaling module with the potential to regulate proteostasis.

COPII proteins exhibit distinct subdomains within each ER exit site for executing their functions.

Secretory proteins are exported from special domains of the endoplasmic reticulum (ER) termed ER exit sites, via COPII-coated carriers. We recently showed that TANGO1 and Sec16 cooperatively organize mammalian ER exit sites for efficient secretion. However, the detailed spatial organization of mammalian ER exit sites is yet to be revealed. Here, we used super-resolution confocal live imaging microscopy (SCLIM) to investigate the localization of endogenous proteins, and we identified domains abundant in transmembrane complexes (TANGO1/cTAGE5/Sec12) juxtaposed to Sec16. Interestingly, this domain can be distinguished from the inner and the outer coats of COPII proteins within each mammalian ER exit site. Cargoes are partially concentrated in the domain for secretion. Our results suggest that mammalian ER exit sites compartmentalize proteins according to their function in COPII vesicle formation.

Not just a cargo receptor for large cargoes; an emerging role of TANGO1 as an organizer of ER exit sites.

Proteins synthesized within the endoplasmic reticulum (ER) are exported from ER exit sites via coat protein complex II (COPII)-coated vesicles. Although the mechanisms of COPII-vesicle formation at the ER exit sites are highly conserved among species, vertebrate cells secrete a wide range of materials, including collagens and chylomicrons, which form bulky structures within the ER that are too large to fit into conventional carriers. Transport ANd Golgi Organization 1 (TANGO1) was initially identified as a cargo receptor for collagens but has been recently rediscovered as an organizer of ER exit sites. We would like to review recent advances in the mechanism of large cargo secretion and organization of ER exit sites through the function of TANGO1.

The binding of TBK1 to STING requires exocytic membrane traffic from the ER.

Stimulator of interferon genes (STING) is essential for the type I interferon and pro-inflammatory responses against DNA pathogens. In response to the presence of cytosolic DNA, STING translocates from the endoplasmic reticulum (ER) to the Golgi, and activates TANK-binding kinase 1 (TBK1), a cytosolic kinase that is essential for the activation of STING-dependent downstream signalling. The organelles where TBK1 binds to STING remain unknown. Here we show that TBK1 binds to STING at the Golgi, not at the ER. Treatment with brefeldin A, an agent to block ER-to-Golgi traffic, or knockdown of Sar1, a small GTPase that regulates coat protein complex II (COP-II)-mediated ER-to-Golgi traffic, inhibited the binding of TBK1 to STING. Endogenous TBK1 was recruited to the Golgi when STING was transported to the Golgi, as shown by immunofluorescence microscopy. STING variants that constitutively induce the type I interferon response were found in patients with autoinflammatory diseases. Even these disease-causative STING variants could not bind to TBK1 when the STING variants were trapped in the ER. These results demonstrate that the Golgi is an organelle at which STING recruits and activates TBK1 for triggering the STING-dependent type I interferon response.

Chemical Events in Oligo(3-methoxythiophene) Coating Solutions and Their Effect on the Goldlike Coating Film Properties.

Metal-free, metal-like lustrous films may find applications in a variety of fields, and a study of the factors affecting their stability is highly desirable. In particular, chemical events occurring in the coating solutions might affect the supramolecular organization of the films and therefore the metal-like luster. Herein, the chemical events occurring in acetonitrile and nitromethane coating solutions of oligo(3-methoxythiophene) and their effect on the optical properties of the films were investigated by X-ray diffraction, UV-vis absorption, and viscosity measurements. In acetonitrile, the oligomers underwent gradual dedoping with time, but only small changes in viscosity were observed. The solution was applied to a glass plate to yield a dark brown film, which turned into a goldlike lustrous film by rubbing. In nitromethane, the supramolecular structure of the oligomers changed with time from the nonaggregated state to π-dimers and then to π-stacks, and the viscosity increased. The properties of the goldlike films prepared from this solution were greatly affected by this chemical event. Remarkably, the π-dimer solution provided the film with the highest specular reflectance, yellowness, greenness, brightness, and crystallinity.

Regulation of the Sar1 GTPase Cycle Is Necessary for Large Cargo Secretion from the Endoplasmic Reticulum.

Proteins synthesized within the endoplasmic reticulum (ER) are transported to the Golgi via coat protein complex II (COPII)-coated vesicles. The formation of COPII-coated vesicles is regulated by the GTPase cycle of Sar1. Activated Sar1 is recruited to ER membranes and forms a pre-budding complex with cargoes and the inner-coat complex. The outer-coat complex then stimulates Sar1 inactivation and completes vesicle formation. The mechanisms of forming transport carriers are well-conserved among species; however, in mammalian cells, several cargo molecules such as collagen, and chylomicrons are too large to be accommodated in conventional COPII-coated vesicles. Thus, special cargo-receptor complexes are required for their export from the ER. cTAGE5/TANGO1 complexes and their isoforms have been identified as cargo receptors for these macromolecules. Recent reports suggest that the cTAGE5/TANGO1 complex interacts with the GEF and the GAP of Sar1 and tightly regulates its GTPase cycle to accomplish large cargo secretion.

CREB3L2-mediated expression of Sec23A/Sec24D is involved in hepatic stellate cell activation through ER-Golgi transport.

Hepatic fibrosis is caused by exaggerated wound healing response to chronic injury, which eventually leads to hepatic cirrhosis. Differentiation of hepatic stellate cells (HSCs) to myofibroblast-like cells by inflammatory cytokines is the critical step in fibrosis. This step is accompanied by enlargement of the endoplasmic reticulum (ER) and Golgi apparatus, suggesting that protein synthesis and secretion are augmented in the activated HSCs. However, the process of rearrangement of secretory organelles and their functions remain to be fully elucidated. Here, we revealed that differentiation alters early secretory gene expression. We observed significant isoform-specific upregulation of the inner coat protein complex II (COPII) components, Sec23A and Sec24D, via the transmembrane bZIP transcription factor, CREB3L2/BBF2H7, during HSC activation. Moreover, knockdown of these components abrogated the activation, suggesting that Sec23A/Sec24D-mediated ER to Golgi trafficking is required for HSC activation.

Remodeling of ER-exit sites initiates a membrane supply pathway for autophagosome biogenesis.

Autophagosomes are double-membrane vesicles generated during autophagy. Biogenesis of the autophagosome requires membrane acquisition from intracellular compartments, the mechanisms of which are unclear. We previously found that a relocation of COPII machinery to the ER-Golgi intermediate compartment (ERGIC) generates ERGIC-derived COPII vesicles which serve as a membrane precursor for the lipidation of LC3, a key membrane component of the autophagosome. Here we employed super-resolution microscopy to show that starvation induces the enlargement of ER-exit sites (ERES) positive for the COPII activator, SEC12, and the remodeled ERES patches along the ERGIC A SEC12 binding protein, CTAGE5, is required for the enlargement of ERES, SEC12 relocation to the ERGIC, and modulates autophagosome biogenesis. Moreover, FIP200, a subunit of the ULK protein kinase complex, facilitates the starvation-induced enlargement of ERES independent of the other subunits of this complex and associates via its C-terminal domain with SEC12. Our data indicate a pathway wherein FIP200 and CTAGE5 facilitate starvation-induced remodeling of the ERES, a prerequisite for the production of COPII vesicles budded from the ERGIC that contribute to autophagosome formation.

TANGO1 recruits Sec16 to coordinately organize ER exit sites for efficient secretion.

Mammalian endoplasmic reticulum (ER) exit sites export a variety of cargo molecules including oversized cargoes such as collagens. However, the mechanisms of their assembly and organization are not fully understood. TANGO1L is characterized as a collagen receptor, but the function of TANGO1S remains to be investigated. Here, we show that direct interaction between both isoforms of TANGO1 and Sec16 is not only important for their correct localization but also critical for the organization of ER exit sites. The depletion of TANGO1 disassembles COPII components as well as membrane-bound ER-resident complexes, resulting in fewer functional ER exit sites and delayed secretion. The ectopically expressed TANGO1 C-terminal domain responsible for Sec16 binding in mitochondria is capable of recruiting Sec16 and other COPII components. Moreover, TANGO1 recruits membrane-bound macromolecular complexes consisting of cTAGE5 and Sec12 to the ER exit sites. These data suggest that mammalian ER exit sites are organized by TANGO1 acting as a scaffold, in cooperation with Sec16 for efficient secretion.