TPC proteins are phosphoinositide- activated sodium-selective ion channels in endosomes and lysosomes.

Mammalian two-pore channel proteins (TPC1, TPC2; TPCN1, TPCN2) encode ion channels in intracellular endosomes and lysosomes and were proposed to mediate endolysosomal calcium release triggered by the second messenger, nicotinic acid adenine dinucleotide phosphate (NAADP). By directly recording TPCs in endolysosomes from wild-type and TPC double-knockout mice, here we show that, in contrast to previous conclusions, TPCs are in fact sodium-selective channels activated by PI(3,5)P(2) and are not activated by NAADP. Moreover, the primary endolysosomal ion is Na(+), not K(+), as had been previously assumed. These findings suggest that the organellar membrane potential may undergo large regulatory changes and may explain the specificity of PI(3,5)P(2) in regulating the fusogenic potential of intracellular organelles.

Biomechanical comparison of traditional anchors to all-suture anchors in a double-row rotator cuff repair cadaver model.

BACKGROUND: To further reduce the invasiveness of arthroscopic rotator cuff repair surgery the all-suture anchor has been developed. The all-suture anchor requires less bone removal and reduces the potential of loose body complications. The all-suture anchor must also have adequate biomechanical strength for the repair to heal. The hypothesis is there is no significant difference in the biomechanical performance of supraspinatus repairs using an all-suture anchor when compared to traditional solid-body suture anchors. METHODS: Using nine shoulders per group, the supraspinatus tendon was dissected from the greater tuberosity. The four different double row repairs tested were (medial row/lateral row): A: ICONIX2/ICONIX2; B: ICONIX2/Stryker ReelX 3.9mm; C: ICONIX2/Stryker ReelX 4.5mm; D: Arthrex BioComposite CorkScrew FT 4.5mm/Arthrex BioComposite SwiveLock 4.75mm. The ICONIX2 was the only all-suture anchor tested. Tendons underwent cyclic loading from 10 to 100N for 500 cycles, followed by load-to-failure. Data was collected at cycles 5, 100, 200, 300, 400, and 500. One-way ANOVA analysis was used to assess significance (P≤0.05). FINDINGS: The anchor combinations tested did not differ significantly in anterior (P>0.4) or posterior (P>0.3) gap formation, construct stiffness (P>0.7), ultimate load (P=0.06), or load to 5mm gap formation (P=0.84). INTERPRETATION: The all-suture anchor demonstrated comparable biomechanical performance in multiple double-row anchor combinations to a combination of traditional solid-body anchors. Thus it may be an attractive option to further reduce the invasiveness of rotator cuff repairs.

A TRP channel in the lysosome regulates large particle phagocytosis via focal exocytosis.

Phagocytosis of large extracellular particles such as apoptotic bodies requires delivery of the intracellular endosomal and lysosomal membranes to form plasmalemmal pseudopods. Here, we identified mucolipin TRP channel 1 (TRPML1) as the key lysosomal Ca2+ channel regulating focal exocytosis and phagosome biogenesis. Both particle ingestion and lysosomal exocytosis are inhibited by synthetic TRPML1 blockers and are defective in macrophages isolated from TRPML1 knockout mice. Furthermore, TRPML1 overexpression and TRPML1 agonists facilitate both lysosomal exocytosis and particle uptake. Using time-lapse confocal imaging and direct patch clamping of phagosomal membranes, we found that particle binding induces lysosomal PI(3,5)P2 elevation to trigger TRPML1-mediated lysosomal Ca2+ release specifically at the site of uptake, rapidly delivering TRPML1-resident lysosomal membranes to nascent phagosomes via lysosomal exocytosis. Thus phagocytic ingestion of large particles activates a phosphoinositide- and Ca2+-dependent exocytosis pathway to provide membranes necessary for pseudopod extension, leading to clearance of senescent and apoptotic cells in vivo.