Bucket lists must be completed during cell death.

Regulated cell death occurs in many forms, including apoptosis, pyroptosis, necroptosis, and NETosis. Most obviously, the purpose of these pathways is to kill the cell. However, many cells need to complete a set of effector programs before they die, which we define as a cellular 'bucket list'. These effector programs are specific to the cell type, and mode and circumstances of death. For example, intestinal epithelial cells need to complete the process of extrusion before they die. Cells use regulatory mechanisms to temporarily prolong their life, including endosomal sorting complex required for transport (ESCRT)- and acid sphingomyelinase (ASM)-driven membrane repair. These allow cells to complete their bucket lists before they die.

Radiation vs surgery for early-stage laryngeal verrucous carcinoma: A population-based propensity score matched-study.

BACKGROUND: Verrucous carcinoma (VC) is a rare variant of squamous cell carcinoma. Although VC is considered radioresistant, concrete evidence for this is absent. METHODS: We obtained data on VC treated with surgery or radiation from the Surveillance, Epidemiology, and End Results database. Treatment selection bias was reduced by propensity score matching. Overall survival (OS) and disease-specific survival (DSS) rates were estimated using the Kaplan-Meier method. Hazard ratios (HRs) were estimated using Cox proportional hazards models. RESULTS: Five-year OS rates in the radiation and surgery groups were 72.7% and 72.0%, respectively (P = 0.111); five-year DSS rates in the same were 86.7% and 88.4%, respectively (P = 0.234). HRs of radiation compared with surgery were 1.68 (95% confidence interval (CI), 0.96-2.95) for OS and 1.95 (95% CI, 0.69-5.53) for DSS. CONCLUSIONS: Similar prognoses were observed in patients with VC treated with radiation and surgery. VC can be treated using radiation.

Caspase-7 activates ASM to repair gasdermin and perforin pores.

Among the caspases that cause regulated cell death, a unique function for caspase-7 has remained elusive. Caspase-3 performs apoptosis, whereas caspase-7 is typically considered an inefficient back-up. Caspase-1 activates gasdermin D pores to lyse the cell; however, caspase-1 also activates caspase-7 for unknown reasons1. Caspases can also trigger cell-type-specific death responses; for example, caspase-1 causes the extrusion of intestinal epithelial cell (IECs) in response to infection with Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium)2,3. Here we show in both organoids and mice that caspase-7-deficient IECs do not complete extrusion. Mechanistically, caspase-7 counteracts gasdermin D pores and preserves cell integrity by cleaving and activating acid sphingomyelinase (ASM), which thereby generates copious amounts of ceramide to enable enhanced membrane repair. This provides time to complete the process of IEC extrusion. In parallel, we also show that caspase-7 and ASM cleavage are required to clear Chromobacterium violaceum and Listeria monocytogenes after perforin-pore-mediated attack by natural killer cells or cytotoxic T lymphocytes, which normally causes apoptosis in infected hepatocytes. Therefore, caspase-7 is not a conventional executioner but instead is a death facilitator that delays pore-driven lysis so that more-specialized processes, such as extrusion or apoptosis, can be completed before cell death. Cells must put their affairs in order before they die.

The AIM2 inflammasome is activated in astrocytes during the late phase of EAE.

Inflammasomes are a class of innate immune signaling platforms that activate in response to an array of cellular damage and pathogens. Inflammasomes promote inflammation under many circumstances to enhance immunity against pathogens and inflammatory responses through their effector cytokines, IL-1ß and IL-18. Multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE), are autoimmune conditions influenced by inflammasomes. Despite work investigating inflammasomes during EAE, little remains known concerning the role of inflammasomes in the central nervous system (CNS) during the disease. Here, we used multiple genetically modified mouse models to monitor activated inflammasomes in situ based on oligomerization of apoptosis-associated speck-like protein containing a CARD (ASC) in the spinal cord. Using inflammasome reporter mice, we found heightened inflammasome activation in astrocytes after the disease peak. In contrast, microglia and CNS-infiltrated myeloid cells had few activated inflammasomes in the CNS during EAE. Astrocyte inflammasome activation during EAE was dependent on absent in melanoma 2 (AIM2), but low IL-1ß release and no significant signs of cell death were found. Thus, the AIM2 inflammasome activation in astrocytes may have a distinct role from traditional inflammasome-mediated inflammation.

Innate Sensors Trigger Regulated Cell Death to Combat Intracellular Infection.

Intracellular pathogens pose a significant threat to animals. In defense, innate immune sensors attempt to detect these pathogens using pattern recognition receptors that either directly detect microbial molecules or indirectly detect their pathogenic activity. These sensors trigger different forms of regulated cell death, including pyroptosis, apoptosis, and necroptosis, which eliminate the infected host cell niche while simultaneously promoting beneficial immune responses. These defenses force intracellular pathogens to evolve strategies to minimize or completely evade the sensors. In this review, we discuss recent advances in our understanding of the cytosolic pattern recognition receptors that drive cell death, including NLRP1, NLRP3, NLRP6, NLRP9, NLRC4, AIM2, IFI16, and ZBP1.

Intraepithelial Lymphocytes Suppress Intestinal Tumor Growth by Cell-to-Cell Contact via CD103/E-Cadherin Signal.

BACKGROUND & AIMS: The reason why small intestinal cancer is rarer than colorectal cancer is not clear. We hypothesized that intraepithelial lymphocytes (IELs), which are enriched in the small intestine, are the closest immune cells to epithelial cells, exclude tumor cells via cell-to-cell contact. METHODS: We developed DPE-green fluorescent protein (DPE-GFP) × adenomatous polyposis coli; multiple intestinal neoplasia (APCmin ) mice, which is a T-cell-reporter mouse with spontaneous intestinal tumors. We visualized the dynamics of IELs in the intestinal tumor microenvironment and the interaction between IELs and epithelial cells, and the roles of cell-to-cell contact in anti-intestinal tumor immunity using a novel in vivo live-imaging system and a novel in vitro co-culture system. RESULTS: In the small intestinal tumor microenvironment, T-cell movement was restricted around blood vessels and the frequency of interaction between IELs and epithelial cells was reduced. Genetic deletion of CD103 decreased the frequency of interaction between IELs and epithelial cells, and increased the number of small intestinal tumors. In the co-culture system, wild-type IELs expanded and infiltrated to intestinal tumor organoids from APCmin mice and reduced the viability of them, which was cell-to-cell contact and CD103 dependent. CONCLUSIONS: The abundance of IELs in the small intestine may contribute to a low number of tumors, although this system may not work in the colon because of the sparseness of IELs. Strategies to increase the number of IELs in the colon or enhance cell-to-cell contact between IELs and epithelial cells may be effective for the prevention of intestinal tumors in patients with a high cancer risk.

Vertical head impulse and caloric are complementary but react opposite to Meniere's disease hydrops.

OBJECTIVES/HYPOTHESIS: Meniere's disease (MD) patients can show normal head impulses despite poor caloric test results. This study aimed to investigate the discrepancy in the vestibulo-ocular reflex (VOR) in MD patients and whether endolymphatic hydrops (EH) influence the VOR. STUDY DESIGN: Prospective, cross-sectional observational study. METHODS: Ninety MD patients were enrolled. Neuro-otological testing, including a video head impulse test (vHIT) of all semicircular canals (SCs), and gadolinium-enhanced inner ear magnetic resonance imaging were performed. The vestibular EH volume was quantitatively evaluated by processing magnetic resonance images. RESULTS: Abnormal vHIT results in MD patients were found most frequently in the posterior (44.4%) SCs, followed by the horizontal (13.3%) and anterior (10%) SCs. Canal paresis (CP) was assessed using the vHIT and the caloric test, and results were not significant when vHIT responses were assessed as CP only using the horizontal SC. The difference in the vestibular EH between the presence and absence of CP was not significant if assessed using the vHIT (P = .5591), but it was statistically different if assessed using the caloric test (P = .0467). CONCLUSIONS: The contradictory reaction of VOR in MD patients may result from the high specificity but low sensitivity of CP in the horizontal vHIT. EH volume in the vestibule affects the caloric response but does not affect the vHIT response. LEVEL OF EVIDENCE: 2b Laryngoscope, 129:1660-1666, 2019.

Room temperature continuous-wave nanolaser diode utilized by ultrahigh-Q few-cell photonic crystal nanocavities.

Few-cell point-defect photonic crystal (PhC) nanocavities (such as LX and H1 type cavities), have several unique characteristics including an ultra-small mode volume (Vm), a small device footprint advantageous for dense integration, and a large mode spacing advantageous for high spontaneous-emission coupling coefficient (ß), which are promising for energy-efficient densely-integratable on-chip laser light sources enhanced by the cavity QED effect. To achieve this goal, a high quality factor (Q) is essential, but conventional few-cell point-defect cavities do not have a sufficiently high Q. Here we adopt a series of modified designs of LX cavities with a buried heterostructure (BH) multi-quantum-well (MQW) active region that can achieve a high Q while maintaining their original advantages and fabricate current-injection laser devices. We have successfully observed continuous-wave (CW) lasing in InP-based L1, L2, L3 and L5 PhC nanocavities at 23°C with a DC current injection lower than 10 µA and a bias voltage lower than 0.9 V. The active volume is ultra-small while maintaining a sufficiently high confinement factor, which is as low as ~10-15 cm3 for a single-cell (L1) nanocavity. This is the first room-temperature current-injection CW lasing from any types of few-cell point-defect PhC nanocavities (LX or H1 types). Our report marks an important step towards realizing a nanolaser diode with a high cavity-QED effect, which is promising for use with on-chip densely integrated laser sources in photonic networks-on-chip combined with CMOS processors.

Wideband slow short-pulse propagation in one-thousand slantingly coupled L3 photonic crystal nanocavities.

Coupled cavities have been used previously to realize on-chip low-dispersion slow-light waveguides, but the bandwidth was usually narrower than 10 nm and the total length was much shorter than 1 mm. Here we report long (0.05-2.5 mm) slow-light coupled cavity waveguides formed by using 50, 200, and 1,000 L3 photonic crystal nanocavities with an optical volume smaller than (λ/n)3, slanted from Γ-K orientation. We demonstrate experimentally the formation of a single-mode wideband coupled cavity mode with a bandwidth of up to 32nm (4THz) in telecom C-band, generated from the ultra-narrow-band (~300 MHz) fundamental mode of each L3 nanocavity, by controlling the cavity array orientation. Thanks to the ultrahigh-Q nanocavity design, coupled cavity waveguides longer than 1 mm exhibited low loss and allowed time-of-flight dispersion measurement over a bandwidth up to 22 nm by propagating a short pulse over 1,000 coupled L3 nanocavities. The highly-dense slanted array of L3 nanocavity demonstrated unprecedentedly high cavity coupling among the nanocavities. The scheme we describe provides controllable planar dispersion-managed waveguides as an alternative to W1-based waveguides on a photonic crystal chip.

Distinct intestinal adaptation for vitamin B12 and bile acid absorption revealed in a new mouse model of massive ileocecal resection.

Ileocecal resection (ICR), one of several types of intestinal resection that results in short bowel syndrome (SBS), causes severe clinical disease in humans. We here describe a mouse model of massive ICR in which 75% of the distal small intestine is removed. We demonstrate that mice underwent 75% ICR show severe clinical signs and high mortality, which may recapitulate severe forms of human SBS, despite an adaptive response throughout the remnant intestine. By using this model, we also investigated whether the epithelium of the remnant intestine shows enhanced expression of factors involved in region-specific functions of the ileum. Cubn mRNA and its protein product, which play an essential role in vitamin B12 absorption in the ileum, are not compensatory up-regulated in any part of the remnant intestine, demonstrating a clear contrast with post-operative up-regulation of genes involved in bile acid absorption. Our study suggests that functional adaptation by phenotypical changes in the intestinal epithelium is not a general feature for nutrient absorption systems that are confined to the ileum. We also propose that the mouse model developed in this study will become a unique system to facilitate studies on SBS with ICR in humans.

Retinol Promotes In Vitro Growth of Proximal Colon Organoids through a Retinoic Acid-Independent Mechanism.

Retinol (ROL), the alcohol form of vitamin A, is known to control cell fate decision of various types of stem cells in the form of its active metabolite, retinoic acid (RA). However, little is known about whether ROL has regulatory effects on colonic stem cells. We examined in this study the effect of ROL on the growth of murine normal colonic cells cultured as organoids. As genes involved in RA synthesis from ROL were differentially expressed along the length of the colon, we tested the effect of ROL on proximal and distal colon organoids separately. We found that organoid forming efficiency and the expression level of Lgr5, a marker gene for colonic stem cells were significantly enhanced by ROL in the proximal colon organoids, but not in the distal ones. Interestingly, neither retinaldehyde (RAL), an intermediate product of the ROL-RA pathway, nor RA exhibited growth promoting effects on the proximal colon organoids, suggesting that ROL-dependent growth enhancement in organoids involves an RA-independent mechanism. This was confirmed by the observation that an inhibitor for RA-mediated gene transcription did not abrogate the effect of ROL on organoids. This novel role of ROL in stem cell maintenance in the proximal colon provides insights into the mechanism of region-specific regulation for colonic stem cell maintenance.

Co-culture with intestinal epithelial organoids allows efficient expansion and motility analysis of intraepithelial lymphocytes.

BACKGROUND: Intraepithelial lymphocytes (IELs) in the intestine play important roles in the regulation of local immune responses. Although their functions have been studied in a variety of animal experiments, in vitro studies on spatiotemporal behaviors of IELs and their interaction with intestinal epithelial cells (IECs) have been hampered due to the lack of a suitable culture system. In this study, we aimed at developing a novel co-culture system of IELs with IECs to investigate dynamic interaction between these two populations of cells in vitro. METHODS: We optimized experimental conditions under which murine IELs can be efficiently maintained with IECs cultured as three-dimensional organoids. We then tested the effect of IL-2, IL-7, and IL-15 on the maintenance of IELs in this co-culture system. By time-lapse imaging, we also examined the dynamic behaviors of IELs. RESULTS: IELs can be expanded with epithelial organoids in the presence of IL-2, IL-7, and IL-15. IELs were efficiently maintained within and outside of organoids showing a ~four-fold increase in both αßT and γδT IELs for a period of 2 weeks. Four-dimensional fluorescent imaging revealed an active, multi-directional movement of IELs along the basolateral surface of IECs, and also their inward or outward migration relative to organoid structures. Cell tracking analysis showed that αßT and γδT IELs shared indistinguishable features with regard to their dynamics. CONCLUSIONS: This novel co-culture method could serve as a unique tool to investigate the motility dynamics of IELs and their temporal and spatial interaction with IECs in vitro.

All-optical switching for 10-Gb/s packet data by using an ultralow-power optical bistability of photonic-crystal nanocavities.

An all-optical packet switching using bistable photonic crystal nanocavity memories was demonstrated for the first time. Nanocavity-waveguide coupling systems were configured for 1 × 1, 1 × 2, and 1 × 3 switches for 10-Gb/s optical packet, and they were all operated with an optical bias power of only a few µW. The power is several magnitudes lower than that of previously reported all-optical packet switches incorporating all-optical memories. A theoretical investigation indicated the optimum design for reducing the power consumption even further, and for realizing a higher data-rate capability and higher extinction. A small footprint and integrability are also features of our switches, which make them attractive for constructing an all-optical packet switching subsystem with a view to realizing optical routing on a chip.

TNFAIP3 promotes survival of CD4 T cells by restricting MTOR and promoting autophagy.

Autophagy plays important roles in metabolism, differentiation, and survival in T cells. TNFAIP3/A20 is a ubiquitin-editing enzyme that is thought to be a negative regulator of autophagy in cell lines. However, the role of TNFAIP3 in autophagy remains unclear. To determine whether TNFAIP3 regulates autophagy in CD4 T cells, we first analyzed Tnfaip3-deficient naïve CD4 T cells in vitro. We demonstrated that Tnfaip3-deficient CD4 T cells exhibited reduced MAP1LC3/LC3 (microtubule-associated protein 1 light chain 3) puncta formation, increased mitochondrial content, and exaggerated reactive oxygen species (ROS) production. These results indicate that TNFAIP3 promotes autophagy after T cell receptor (TCR) stimulation in CD4 T cells. We then investigated the mechanism by which TNFAIP3 promotes autophagy signaling. We found that TNFAIP3 bound to the MTOR (mechanistic target of rapamycin) complex and that Tnfaip3-deficient cells displayed enhanced ubiquitination of the MTOR complex and MTOR activity. To confirm the effects of enhanced MTOR activity in Tnfaip3-deficient cells, we analyzed cell survival following treatment with Torin1, an MTOR inhibitor. Tnfaip3-deficient CD4 T cells exhibited fewer cell numbers than the control cells in vitro and in vivo. In addition, the impaired survival of Tnfaip3-deficient cells was ameliorated with Torin1 treatment in vitro and in vivo. The effect of Torin1 was abolished by Atg5 deficiency. Thus, enhanced MTOR activity regulates the survival of Tnfaip3-deficient CD4 T cells. Taken together, our findings illustrate that TNFAIP3 restricts MTOR signaling and promotes autophagy, providing new insight into the manner in which MTOR and autophagy regulate survival in CD4 T cells.

Systematic hole-shifting of L-type nanocavity with an ultrahigh Q factor.

We report simple systematic hole-shifting rules applicable to any Lx (x:2,3,4,5,…) nanocavity. The rules specify six sets of holes to be tuned with only two or three shift parameters. While keeping the same cavity wavelength and nearly the same mode volume, the new rule increases the Q factor by nearly one order of magnitude compared with an edge-hole-shifted Lx nanocavity. The Q factor of the high-order mode is also greatly increased. This merit is obvious from the maximum experimental Q factors of over 500,000 at L2 and of over 1,000,000 at L3, L4, and L5 achieved in Si photonic crystals.

Compact 1D-silicon photonic crystal electro-optic modulator operating with ultra-low switching voltage and energy.

We demonstrate a small foot print (600 nm wide) 1D silicon photonic crystal electro-optic modulator operating with only a 50 mV swing voltage and 0.1 fJ/bit switching energy at GHz speeds, which are the lowest values ever reported for a silicon electro-optic modulator. A 3 dB extinction ratio is demonstrated with an ultra-low 50 mV swing voltage with a total device energy consumption of 42.8 fJ/bit, which is dominated by the state holding energy. The total energy consumption is reduced to 14.65 fJ/bit for a 300 mV swing voltage while still keeping the switching energy at less than 2 fJ/bit. Under optimum voltage conditions, the device operates with a maximum speed of 3 Gbps with 8 dB extinction ratio, which rises to 11 dB for a 1 Gbps modulation speed.

Small intestinal stem cell identity is maintained with functional Paneth cells in heterotopically grafted epithelium onto the colon.

To develop stem cell therapy for small intestinal (SI) diseases, it is essential to determine whether SI stem cells in culture retain their tissue regeneration capabilities. By using a heterotopic transplantation approach, we show that cultured murine SI epithelial organoids are able to reconstitute self-renewing epithelia in the colon. When stably integrated, the SI-derived grafts show many features unique only to the SI but distinct from the colonic epithelium. Our study provides evidence that cultured adult SI stem cells could be a source for cell therapy of intestinal diseases, maintaining their identity along the gastrointestinal tract through an epithelium-intrinsic mechanism.

25-channel all-optical gate switches realized by integrating silicon photonic crystal nanocavities.

Silicon-based photonic crystal nanocavities with different lattice pitches were monolithically integrated with a total length of only 200 µm, and were operated as a multi-channel all-optical switch with a large processing density of 42 Tb/s/mm2. A pump light and a signal light were assigned to two cavity modes in each cavity, and in this way all-optical gate switching was achieved in a 25 channel resonant dip with an energy cost in the femtojoule regime. We also demonstrated a wavelength-division multiplexing operation by selecting three neighboring channels, and thus achieved gate switching without inter-channel optical crosstalk. As far as we know, this was the first demonstration of a many-channel all-optical switch that can handle an optical signal with bit-by-bit Gb/s repetition in an integrated photonic crystal chip.

InGaAs nano-photodetectors based on photonic crystal waveguide including ultracompact buried heterostructure.

Ultrasmall InGaAs photodetectors based on a photonic crystal waveguide with a buried heterostructure (BH) were demonstrated for the first time. A sufficiently high DC responsivity of ~1 A/W was achieved for the 3.4-µm-long detector. The dynamic response revealed a 3-dB bandwidth of 6 GHz and a 10-Gb/s eye pattern. These results were thanks to the strong confinement of both photons and carriers in a small BH and will pave the way for unprecedented nano-photodetectors with a high quantum efficiency and small capacitance. Our device potentially has an ultrasmall junction capacitance of much less than 1 fF and may enable us to eliminate electrical amplifiers for future optical receivers and subsequent ultralow-power optical links on a chip.