Acid-Labile Traceless Click Linker for Protein Transduction

Intracellular delivery of active proteins presents an interesting approach in research and therapy. We created a protein transduction shuttle based on a new traceless click linker that combines the advantages of click reactions with implementation of reversible pH-sensitive bonds. The azidomethyl-methylmaleic anhydride (AzMMMan) linker was found compatible with different click chemistries, demonstrated in bioreversible protein modification with dyes, polyethylene glycol, or a transduction carrier. Linkages were stable at physiological pH but reversible at the mild acidic pH of endosomes or lysosomes. We show that pH-reversible attachment of a defined endosome-destabilizing three-arm oligo(ethane amino)amide carrier generates an effective shuttle for protein delivery. The cargo protein nlsEGFP, when coupled via the traceless AzMMMan linker, experiences efficient cellular uptake and endosomal escape into the cytosol, followed by import into the nucleus. In contrast, irreversible linkage to the same shuttle hampers nuclear delivery of nlsEGFP which after uptake remains trapped in the cytosol. Successful intracellular delivery of bioactive ß-galactosidase as a model enzyme was also demonstrated using the pH-controlled shuttle system

Integrated frequency comb source of heralded single photons

We report an integrated photon pair source based on a CMOS-compatible microring resonator that generates multiple, simultaneous, and independent photon pairs at different wavelengths in a frequency comb compatible with fiber communication wavelength division multiplexing channels (200 GHz channel separation) and with a linewidth that is compatible with quantum memories (110 MHz). It operates in a self-locked pump configuration, avoiding the need for active stabilization, making it extremely robust even at very low power levels

Chemical chaperone treatment reduces intracellular accumulation of mutant collagen IV and ameliorates the cellular phenotype of a COL4A2 mutation that causes haemorrhagic stroke

Haemorrhagic stroke accounts for approximately 20% of stroke cases and porencephaly is a clinical consequence of perinatal cerebral haemorrhaging. Here we report the identification of a novel dominant G702D mutation in the collagen domain of COL4A2 (collagen IV alpha chain 2) in a family displaying porencephaly with reduced penetrance. COL4A2 is the obligatory protein partner of COL4A1 but in contrast to most COL4A1 mutations, the COL4A2 mutation does not lead to eye or kidney disease. Analysis of dermal biopsies from patient and his unaffected father, who also carries the mutation, revealed that both display basement membrane (BM) defects. Intriguingly, defective collagen IV incorporation into the dermal BM was only observed in the patient and was associated with endoplasmic reticulum (ER) retention of COL4A2 in primary dermal fibroblasts. This intracellular accumulation led to ER-stress, unfolded protein response activation, reduced cell proliferation and increased apoptosis. Interestingly, absence of ER retention of COL4A2 and ER-stress in cells from the unaffected father indicate that accumulation and/or clearance of mutant COL4A2 from the ER may be a critical modifier for disease development. Our analysis also revealed that mutant collagen IV is degraded via the proteasome. Importantly, treatment of patient cells with a chemical chaperone decreased intracellular COL4A2, ER-stress and apoptosis, demonstrating that reducing intracellular collagen accumulation can ameliorate the cellular phenotype of COL4A2 mutations. Importantly, these data highlight that manipulation of chaperone levels, intracellular collagen accumulation and ER-stress are potential therapeutic options for collagen IV diseases including haemorrhagic stroke

Clinicopathological Profile and Surgical Treatment of Abdominal Tuberculosis: A Single Centre Experience in Northwestern Tanzania.

Abdominal tuberculosis continues to be a major public health problem worldwide and poses diagnostic and therapeutic challenges to general surgeons practicing in resource-limited countries. This study was conducted to describe the clinicopathological profile and outcome of surgical treatment of abdominal tuberculosis in our setting and compare with what is described in literature. A prospective descriptive study of patients who presented with abdominal tuberculosis was conducted at Bugando Medical Centre (BMC) in northwestern Tanzania from January 2006 to February 2012. Ethical approval to conduct the study was obtained from relevant authorities. Statistical data analysis was performed using SPSS version 17.0. Out of 256 patients enrolled in the study, males outnumbered females. The median age was 28 years (range = 16-68 years). The majority of patients (77.3%) had primary abdominal tuberculosis. A total of 127 (49.6%) patients presented with intestinal obstruction, 106 (41.4%) with peritonitis, 17 (6.6%) with abdominal masses and 6 (2.3%) patients with multiple fistulae in ano. Forty-eight (18.8%) patients were HIV positive. A total of 212 (82.8%) patients underwent surgical treatment for abdominal tuberculosis. Bands /adhesions (58.5%) were the most common operative findings. Ileo-caecal region was the most common bowel involved in 122 (57.5%) patients. Release of adhesions and bands was the most frequent surgical procedure performed in 58.5% of cases. Complication and mortality rates were 29.7% and 18.8% respectively. The overall median length of hospital stay was 32 days and was significantly longer in patients with complications (p < 0.001). Advanced age (age ≥ 65 years), co-morbid illness, late presentation, HIV positivity and CD4+ count < 200 cells/μl were statistically significantly associated with mortality (p < 0.0001). The follow up of patients were generally poor as only 37.5% of patients were available for follow up at twelve months after discharge. Abdominal tuberculosis constitutes a major public health problem in our environment and presents a diagnostic challenge requiring a high index of clinical suspicion. Early diagnosis, early anti-tuberculous therapy and surgical treatment of the associated complications are essential for survival

Swirl Flow Bioreactor coupled with Cu-alginate beads: A system for the eradication of Coliform and Escherichia coli from biological effluents.

It is estimated that approximately 1.1 billion people globally drink unsafe water. We previously reported both a novel copper-alginate bead, which quickly reduces pathogen loading in waste streams and the incorporation of these beads into a novel swirl flow bioreactor (SFB), of low capital and running costs and of simple construction from commercially available plumbing pipes and fittings. The purpose of the present study was to trial this system for pathogen reduction in waste streams from an operating Dewats system in Hinjewadi, Pune, India and in both simulated and real waste streams in Seattle, Washington, USA. The trials in India, showed a complete inactivation of coliforms in the discharged effluent (Mean Log removal Value (MLRV) = 3.51), accompanied by a total inactivation of E. coli with a MLRV of 1.95. The secondary clarifier effluent also showed a 4.38 MLRV in viable coliforms during treatment. However, the system was slightly less effective in reducing E. coli viability, with a MLRV of 1.80. The trials in Seattle also demonstrated the efficacy of the system in the reduction of viable bacteria, with a LRV of 5.67 observed of viable Raoultella terrigena cells (100%)

The Influence of the effect of solute on the thermodynamic driving force on grain refinement of Al alloys

Grain refinement is known to be strongly affected by the solute in cast alloys. Addition of some solute can reduce grain size considerably while others have a limited effect. This is usually attributed to the constitutional supercooling which is quantified by the growth restriction factor, Q. However, one factor that has not been considered is whether different solutes have differing effects on the thermodynamic driving force for solidification. This paper reveals that addition of solute reduces the driving force for solidification for a given undercooling, and that for a particular Q value, it is reduced more substantially when adding eutectic-forming solutes than peritectic-forming elements. Therefore, compared with the eutectic-forming solutes, addition of peritectic-forming solutes into Al alloys not only possesses a higher initial nucleation rate resulted from the larger thermodynamic driving force for solidification, but also promotes nucleation within the constitutionally supercooled zone during growth. As subsequent nucleation can occur at smaller constitutional supercoolings for peritectic-forming elements, a smaller grain size is thus produced. The very small constitutional supercooling required to trigger subsequent nucleation in alloys containing Ti is considered as a major contributor to its extraordinary grain refining efficiency in cast Al alloys even without the deliberate addition of inoculants.The Australian Research Council (ARC DP10955737)

4-Phenylbutyric acid treatment rescues trafficking and processing of a mutant surfactant protein C

Mutations in the SFTPC gene, encoding surfactant protein–C (SP-C), are associated with interstitial lung disease (ILD). Knowledge of the intracellular fate of mutant SP-C is essential in the design of therapies to correct trafficking/processing of the proprotein, and to prevent the formation of cytotoxic aggregates. We assessed the potential of a chemical chaperone to correct the trafficking and processing of three disease-associated mutant SP-C proteins. HEK293 cells were stably transfected with wild-type (SP-C(WT)) or mutant (SP-C(L188Q), SP-C(Δexon4), or SP-C(I73T)) SP-C, and cell lines with a similar expression of SP-C mRNA were identified. The effects of the chemical chaperone 4-phenylbutyric acid (PBA) and lysosomotropic drugs on intracellular trafficking to the endolysosomal pathway and the subsequent conversion of SP-C proprotein to mature peptide were assessed. Despite comparable SP-C mRNA expression, proprotein concentrations varied greatly: SP-C(I73T) was more abundant than SP-C(WT) and was localized to the cell surface, whereas SP-C(Δexon4) was barely detectable. In contrast, SP-C(L188Q) and SP-C(WT) proprotein concentrations were comparable, and a small amount of SP-C(L188Q) was localized to the endolysosomal pathway. PBA treatment restored the trafficking and processing of SP-C(L188Q) to SP-C(WT) concentrations, but did not correct the mistrafficking of SP-C(I73T) or rescue SP-C(Δexon4). PBA treatment also promoted the aggregation of SP-C proproteins, including SP-C(L188Q). This study provides proof of the principle that a chemical chaperone can correct the mistrafficking and processing of a disease-associated mutant SP-C proprotein