Reconstruction of the palatal aponeurosis with autogenous fascia lata in secondary radical intravelar veloplasty: a new method

Velopharyngeal insufficiency in cleft patients with muscular insufficiency detected by nasendoscopy is commonly treated by secondary radical intravelar veloplasty, in which the palatal muscles are reoriented and positioned backwards. The dead space between the retro-displaced musculature and the posterior borders of the palatal bone remains problematic. Postoperatively, the surgically achieved lengthening of the soft palate often diminishes due to scar tissue formation in the dead space, leading to reattachment of the reoriented muscles to the palatal bone and to decreased mobility of the soft palate. To avoid this, the dead space should be restored by a structure imitating the function of the missing palatal aponeurosis. The entire dead space was covered using a double layer of autogenous fascia lata harvested from the lateral thigh, which should allow sufficient and permanent sliding of the retro-positioned musculature. A clinical case of a 9-year-old boy who underwent the operation is reported. Postoperatively, marked functional improvements were observable in speech assessment, nasendoscopy and nasometry. The case reported here suggests that the restoration of the dead space may be beneficial for effective secondary palatal repair. Fascia lata seems to be a suitable graft for this purpose.

Establishment and expansion of Lake Malawi rock fish populations after a dramatic Late Pleistocene lake level rise

Major environmental events that fragment populations among multiple island habitats have potential to drive large-scale episodes of speciation and adaptive radiation. A recent palaeolimnological study of sediment cores indicated that Lake Malawi underwent major climate-driven desiccation events 75 000-135 000 years ago that lowered the water level to at least 580 m below the present state and severely reduced surface area. After this period, lake levels rose and stabilized, creating multiple discontinuous littoral rocky habitats. Here, we present evidence supporting the hypothesis that establishment and expansion of isolated philopatric rock cichlid populations occurred after this rise and stabilization of lake level. We studied the Pseudotropheus (Maylandia) species complex, a group with both allopatric and sympatric populations that differ in male nuptial colour traits and tend to mate assortatively. Using coalescent analyses based on mitochondrial DNA, we found evidence that populations throughout the lake started to expand and accumulate genetic diversity after the lake level rise. Moreover, most haplotypes were geographically restricted, and the greatest genetic similarities were typically among sympatric or neighbouring populations. This is indicative of limited dispersal and establishment of assortative mating among populations following the lake level rise. Together, this evidence is compatible with a single large-scale environmental event being central to evolution of spatial patterns of genetic and species diversity in P. (Maylandia) and perhaps other Lake Malawi rock cichlids. Equivalent climate-driven pulses of habitat formation and fragmentation may similarly have contributed to observed rapid and punctuated cladogenesis in other adaptive radiations.

Characterization and optimization of antigen-specific T cell responses during ex vivo expansion of melanoma tumor-infiltrating lymphocytes

Treatment of metastatic melanoma with tumor reactive T cells (adoptive T cell therapy, ACT) is a promising approach associated with a high clinical response rate. However, further optimization of this treatment modality is required to increase the clinical response after this therapy. ACT in melanoma involves an initial phase (pre-REP) of tumor-infiltrating lymphocyte (TIL) expansion ex vivo from tumor isolates followed by a second phase, “rapid expansion protocol” (REP) generating the billions of cells used as the TIL infusion product. The main question addressed in this thesis was how the currently used REP affected the responsiveness of the CD8+ T cells to defined melanoma antigens. We hypothesized that the REP drives the TIL to further differentiate and become hyporesponsive to antigen restimulation, therefore, proper cytokine treatment or other ways to expand TIL is required to improve upon this outcome. We evaluated the response of CD8+ TIL to melanoma antigen restimulation using MART-1 peptide-pulsed mature DC in vitro. Post-REP TILs were mostly hypo-responsive with poor proliferation and higher apoptosis. Phenotypic analysis revealed that the expression of CD28 was significantly reduced in post-REP TILs. By sorting experiment and microarray analysis, we confirmed that the few CD28+ post-REP TILs had superior survival capacity and proliferated after restimulation. We then went on to investigate methods to maintain CD28 expression during the REP and improve TIL responsiveness. Firstly, IL-15 and IL-21 were found to synergize in maintaining TIL CD28 expression and antigenic responsiveness during REP. Secondly, we found IL-15 was superior as compared to IL-2 in supporting the long-term expansion of antigen-specific CD8+ TIL after restimulation. These results suggest that current expansion protocols used for adoptive T-cell therapy in melanoma yield largely hyporesponsive products containing CD8+ T cells unable to respond in vivo to re-stimulation with antigen. A modification of our current approaches by using IL-15+IL-21 as supporting cytokines in the REP, or/and administration of IL-15 instead of IL-2 after TIL infusion, may enhance the anti-tumor efficacy and long-term persistence of infused T cells in vivo.

Vegetation responses to rapid warming and to minor climatic fluctuations during the Late-Glacial Interstadial (GI-1) at Gerzensee (Switzerland)

High-resolution pollen analyses made on the same samples on which the ratios of oxygen isotopes were measured that provided the time scale and a temperature proxy after correlation to NorthGRIP. (1) A primary succession: The vegetation responded to the rapid rise of temperatures around 14,685 yr BP, with a primary succession on a decadal to centennial time scale. The succession between ca 15,600 and 13,000 yr BP included: (1.1.) The replacement of shrub-tundra by woodland of Juniperus and tree birch (around 14,665 yr BP) (1.2.) The response of Juniperus pollen to the shift in oxygen isotopes in less than 20 yr, (1.3.) A sequence of population increases of Hippophaë rhamnoides (ca 14,600 yr BP), Salix spp. (ca 14,600 yr BP), Betula trees (ca.14,480 yr BP), Populus cf. tremula (ca. 14,300 yr BP), and Pinus cf. sylvestris (ca. 13,830 yr BP). (2) Biological processes: Plants responded to the rapid increase of summer temperatures on all organisational levels: (2.1) Individuals may have produced more pollen (e.g. Juniperus); (2.2) Populations increased or decreased (e.g. Juniperus, Betula, later Pinus), and (2.3) Populations changed their biogeographical range and may show migrational lags. (2.4) Plant communities changed in their composition because the species pools changed through immigration and (local) extinction. Some plant communities may have been without modern analogue.These mechanisms require increasing amounts of time. (2.5) Processes on the level of ecosystems, with species interactions, may involve various time scales. Besides competition and facilitation, nitrogen fixation is discussed. (3) The minor fluctuations of temperature during the Late-Glacial Interstadial, which are recorded in δ18O, resulted in only very minor changes in pollen during the Aegelsee Oscillation (Older Dryas biozone, GI-1d) and the Gerzensee Oscillation (GI-1b). (4) Biodiversity: The afforestation at the onset of Bølling coincided with a gradual increase of taxonomic diversity up to the time of the major Pinus expansion.

Contemporary ecotypic divergence during a recent range expansion was facilitated by adaptive introgression

Although rapid phenotypic evolution during range expansion associated with colonization of contrasting habitats has been documented in several taxa, the evolutionary mechanisms that underlie such phenotypic divergence have less often been investigated. A strong candidate for rapid ecotype formation within an invaded range is the three-spine stickleback in the Lake Geneva region of central Europe. Since its introduction only about 140 years ago, it has undergone a significant expansion of its range and its niche, now forming phenotypically differentiated parapatric ecotypes that occupy either the pelagic zone of the large lake or small inlet streams, respectively. By comparing museum collections from different times with contemporary population samples, we here reconstruct the evolution of parapatric phenotypic divergence through time. Using genetic data from modern samples, we infer the underlying invasion history. We find that parapatric habitat-dependent phenotypic divergence between the lake and stream was already present in the first half of the twentieth century, but the magnitude of differentiation increased through time, particularly in antipredator defence traits. This suggests that divergent selection between the habitats occurred and was stable through much of the time since colonization. Recently, increased phenotypic differentiation in antipredator defence traits likely results from habitat-dependent selection on alleles that arrived through introgression from a distantly related lineage from outside the Lake Geneva region. This illustrates how hybridization can quickly promote phenotypic divergence in a system where adaptation from standing genetic variation was constrained.

Rapid divergence in physiological and life-history traits between northern and southern populations of the British introduced neo-species, Senecio squalidus

Alien plants provide a unique opportunity to study evolution in novel environments, but relatively little is known about the extent to which they become locally adapted to different environments across their new range. Here, we compare northern and southern populations of the introduced species Senecio squalidus in Britain; S. squalidus has been in southern Britain for approximately 200 years and reached Scotland only about 50 years ago. We conducted common garden experiments at sites in the north and south of the species’ range in Britain. We also conducted glasshouse and growth chamber experiments to test the hypothesis that southern genotypes flower later, are more drought-tolerant, germinate and establish better at warmer temperatures, and are less sensitive to cold stress than their more northern counterparts. Results from the common garden experiments are largely consistent with the hypothesis of rapid adaptive divergence of populations of the species within the introduced range, with genotypes typically showing a home-site advantage. Results from the glasshouse and growth chamber experiments demonstrate adaptive divergence in ability to tolerate drought stress and high temperatures, as well as in phenology. In particular, southern genotypes were more tolerant of dry conditions and high temperatures and they flowered later than northern genotypes. Our results show that rapid local adaptation can occur in alien species, and they have implications for our understanding of the ecological genetics of range expansion of introduced weeds.

Biogeographic range expansion into South America by Coccidioides immitis mirrors New World patterns of human migration

Long-distance population dispersal leaves its characteristic signature in genomes, namely, reduced diversity and increased linkage between genetic markers. This signature enables historical patterns of range expansion to be traced. Herein, we use microsatellite loci from the human pathogen Coccidioides immitis to show that genetic diversity in this fungus is geographically partitioned throughout North America. In contrast, analyses of South American C. immitis show that this population is genetically depauperate and was founded from a single North American population centered in Texas. Variances of allele distributions show that South American C. immitis have undergone rapid population growth, consistent with an epidemic increase in postcolonization population size. Herein, we estimate the introduction into South America to have occurred within the last 9,000–140,000 years. This range increase parallels that of Homo sapiens. Because of known associations between Amerindians and this fungus, we suggest that the colonization of South America by C. immitis represents a relatively recent and rapid codispersal of a host and its pathogen.

Rapid Regulation by Acid pH of Cell Wall Adjustment and Leaf Growth in Maize Plants Responding to Reversal of Water Stress1

The role of acid secretion in regulating short-term changes in growth rate and wall extensibility was investigated in emerging first leaves of intact, water-stressed maize (Zea mays L.) seedlings. A novel approach was used to measure leaf responses to injection of water or solutions containing potential regulators of growth. Both leaf elongation and wall extensibility, as measured with a whole-plant creep extensiometer, increased dramatically within minutes of injecting water, 0.5 mm phosphate, or strong (50 mm) buffer solutions with pH ≤ 5.0 into the cell-elongation zone of water-stressed leaves. In contrast, injecting buffer solutions at pH ≥ 5.5 inhibited these fast responses. Solutions containing 0.5 mm orthovanadate or erythrosin B to inhibit wall acidification by plasma membrane H+-ATPases were also inhibitory. Thus, cell wall extensibility and leaf growth in water-stressed plants remained inhibited, despite the increased availability of (injected) water when accompanying increases in acid-induced wall loosening were prevented. However, growth was stimulated when pH 4.5 buffers were included with the vanadate injections. These findings suggest that increasing the availability of water to expanding cells in water-stressed leaves signals rapid increases in outward proton pumping by plasma membrane H+-ATPases. Resultant increases in cell wall extensibility participate in the regulation of water uptake, cell expansion, and leaf growth.

The heart communicates with the kidney exclusively through the guanylyl cyclase-A receptor: acute handling of sodium and water in response to volume expansion.

Disruption of guanylyl cyclase-A (GC-A) results in mice displaying an elevated blood pressure, which is not altered by high or low dietary salt. However, atrial natriuretic peptide (ANP), a proposed ligand for GC-A, has been suggested as critical for the maintenance of normal blood pressure during high salt intake. In this report, we show that infusion of ANP results in substantial natriuresis and diuresis in wild-type mice but fails to cause significant changes in sodium excretion or urine output in GC-A-deficient mice. ANP, therefore, appears to signal through GC-A in the kidney. Other natriuretic/diuretic factors could be released from the heart. Therefore, acute volume expansion was used as a means to cause release of granules from the atrium of the heart. That granule release occurred was confirmed by measurements of plasma ANP concentrations, which were markedly elevated in both wild-type and GC-A-null mice. After volume expansion, urine output as well as urinary sodium and cyclic GMP excretion increased rapidly and markedly in wild-type mice, but the rapid increases were abolished in GC-A-deficient animals. These results strongly suggest that natriuretic/diuretic factors released from the heart function exclusively through GC-A.

Self-renewal of primitive human hematopoietic cells (long-term-culture-initiating cells) in vitro and their expansion in defined medium.

A major goal of experimental and clinical hematology is the identification of mechanisms and conditions that support the expansion of transplantable hematopoietic stem cells. In normal marrow, such cells appear to be identical to (or represent a subset of) a population referred to as long-term-culture-initiating cells (LTC-ICs) so-named because of their ability to produce colony-forming cell (CFC) progeny for > or = 5 weeks when cocultured with stromal fibroblasts. Some expansion of LTC-ICs in vitro has recently been described, but identification of the factors required and whether LTC-IC self-renewal divisions are involved have remained unresolved issues. To address these issues, we examined the maintenance and/or generation of LTC-ICs from single CD34+ CD38- cells cultured for variable periods under different culture conditions. Analysis of the progeny obtained from cultures containing a feeder layer of murine fibroblasts engineered to produce steel factor, interleukin (IL)-3, and granulocyte colony-stimulating factor showed that approximately 20% of the input LTC-ICs (representing approximately 2% of the original CD34+ CD38- cells) executed self-renewal divisions within a 6-week period. Incubation of the same CD34+ CD38- starting populations as single cells in a defined (serum free) liquid medium supplemented with Flt-3 ligand, steel factor, IL-3, IL-6, granulocyte colony-stimulating factor, and nerve growth factor resulted in the proliferation of initial cells to produce clones of from 4 to 1000 cells within 10 days, approximately 40% of which included > or = 1 LTC-IC. In contrast, in similar cultures containing methylcellulose, input LTC-ICs appeared to persist but not divide. Overall the LTC-IC expansion in the liquid cultures was 30-fold in the first 10 days and 50-fold by the end of another 1-3 weeks. Documentation of human LTC-IC self-renewal in vitro and identification of defined conditions that permit their extensive and rapid amplification should facilitate analysis of the molecular mechanisms underlying these processes and their exploitation for a variety of therapeutic applications.

Oxytocin mediates atrial natriuretic peptide release and natriuresis after volume expansion in the rat.

Our previous studies have shown that stimulation of the anterior ventral third ventricular region increases atrial natriuretic peptide (ANP) release, whereas lesions of this structure, the median eminence, or removal of the neural lobe of the pituitary block ANP release induced by blood volume expansion (BVE). These results indicate that participation of the central nervous system is crucial in these responses, possibly through mediation by neurohypophysial hormones. In the present research we investigated the possible role of oxytocin, one of the two principal neurohypophysial hormones, in the mediation of ANP release. Oxytocin (1-10 nmol) injected i.p. caused significant, dose-dependent increases in urinary osmolality, natriuresis, and kaliuresis. A delayed antidiuretic effect was also observed. Plasma ANP concentrations increased nearly 4-fold (P < 0.01) 20 min after i.p. oxytocin (10 nmol), but there was no change in plasma ANP values in control rats. When oxytocin (1 or 10 nmol) was injected i.v., it also induced a dose-related increase in plasma ANP at 5 min (P < 0.001). BVE by intra-atrial injection of isotonic saline induced a rapid (5 min postinjection) increase in plasma oxytocin and ANP concentrations and a concomitant decrease in plasma arginine vasopressin concentration. Results were similar with hypertonic volume expansion, except that this induced a transient (5 min) increase in plasma arginine vasopressin. The findings are consistent with the hypothesis that baroreceptor activation of the central nervous system by BVE stimulates the release of oxytocin from the neurohypophysis. This oxytocin then circulates to the right atrium to induce release of ANP, which circulates to the kidney and induces natriuresis and diuresis, which restore body fluid volume to normal levels.

Expansion-based algorithms for finding single pair shortest path on surface

Finding single pair shortest paths on surface is a fundamental problem in various domains, like Geographic Information Systems (GIS) 3D applications, robotic path planning system, and surface nearest neighbor query in spatial database, etc. Currently, to solve the problem, existing algorithms must traverse the entire polyhedral surface. With the rapid advance in areas like Global Positioning System (CPS), Computer Aided Design (CAD) systems and laser range scanner, surface models axe becoming more and more complex. It is not uncommon that a surface model contains millions of polygons. The single pair shortest path problem is getting harder and harder to solve. Based on the observation that the single pair shortest path is in the locality, we propose in this paper efficient methods by excluding part of the surface model without considering them in the search process. Three novel expansion-based algorithms are proposed, namely, Naive algorithm, Rectangle-based Algorithm and Ellipse-based Algorithm. Each algorithm uses a two-step approach to find the shortest path. (1) compute an initial local path. (2) use the value of this initial path to select a search region, in which the global shortest path exists. The search process terminates once the global optimum criteria are satisfied. By reducing the searching region, the performance is improved dramatically in most cases.

Tissue-specific bioscaffolds for adipose-derived stem/stromal cell expansion and delivery

Decellularized adipose tissue (DAT) is a promising biomaterial for soft tissue regeneration, and it provides a highly conducive microenvironment for human adipose-derived stem/stromal cell (ASC) attachment, proliferation, and adipogenesis. This thesis focused on developing techniques to fabricate 3-D bioscaffolds from enzymatically-digested DAT as platforms for ASC culture and delivery in adipose tissue engineering and large-scale ASC expansion. Initial work investigated chemically crosslinked microcarriers fabricated from pepsin-digested DAT as injectable adipo-inductive substrates for ASCs. DAT microcarriers highly supported ASC adipogenesis compared to gelatin microcarriers in a CELLSPIN system, as confirmed by glycerol-3-phosphate dehydrogenase (GPDH) enzyme activity, lipid accumulation, and endpoint RT-PCR. ASCs cultured on DAT microcarriers in proliferation medium also had elevated PPARγ, C/EBPα, and LPL expression which suggested adipo-inductive properties. In vivo testing of the DAT microcarriers exhibited stable volume retention and enhanced cellular infiltration, tissue remodeling, and angiogenesis. Building from this work, non-chemically crosslinked porous foams and bead foams were fabricated from α-amylase-digested DAT for soft tissue regeneration. Foams were stable and strongly supported ASC adipogenesis based on GPDH activity and endpoint RT-PCR. PPARγ, C/EBPα, and LPL expression in ASCs cultured on the foams in proliferation media indicated adipo-inductive properties. Foams with Young’s moduli similar to human fat also influenced ASC adipogenesis by enhanced GPDH activity. In vivo adipogenesis accompanied by a potent angiogenic response and rapid resorption showed their potential use in wound healing applications. Finally, non-chemically crosslinked porous microcarriers synthesized from α-amylase-digested DAT were investigated for ASC expansion. DAT microcarriers remained stable in culture and supported significantly higher ASC proliferation compared to Cultispher-S microcarriers in a CELLSPIN system. ASC immunophenotype was preserved for all expanded groups, with reduced adhesion marker expression under dynamic conditions. DAT microcarrier expansion upregulated ASC expression of early adipogenic (PPARγ, LPL) and chondrogenic (COMP) markers without inducing a mature phenotype. DAT microcarrier expanded ASCs also showed similar levels of adipogenesis and osteogenesis compared to Cultispher-S despite a significantly higher population fold-change, and had the highest level of chondrogenesis among all groups. This study demonstrates the promising use of DAT microcarriers as a clinically relevant strategy for ASC expansion while maintaining multilineage differentiation capacity.

Rapid emergence of multidrug resistant, H58-lineage Salmonella typhi in Blantyre, Malawi

INTRODUCTION: Between 1998 and 2010, S. Typhi was an uncommon cause of bloodstream infection (BSI) in Blantyre, Malawi and it was usually susceptible to first-line antimicrobial therapy. In 2011 an increase in a multidrug resistant (MDR) strain was detected through routine bacteriological surveillance conducted at Queen Elizabeth Central Hospital (QECH).

METHODS: Longitudinal trends in culture-confirmed Typhoid admissions at QECH were described between 1998-2014. A retrospective review of patient cases notes was conducted, focusing on clinical presentation, prevalence of HIV and case-fatality. Isolates of S. Typhi were sequenced and the phylogeny of Typhoid in Blantyre was reconstructed and placed in a global context.

RESULTS: Between 1998-2010, there were a mean of 14 microbiological diagnoses of Typhoid/year at QECH, of which 6.8% were MDR. This increased to 67 in 2011 and 782 in 2014 at which time 97% were MDR. The disease predominantly affected children and young adults (median age 11 [IQR 6-21] in 2014). The prevalence of HIV in adult patients was 16.7% [8/48], similar to that of the general population (17.8%). Overall, the case fatality rate was 2.5% (3/94). Complications included anaemia, myocarditis, pneumonia and intestinal perforation. 112 isolates were sequenced and the phylogeny demonstrated the introduction and clonal expansion of the H58 lineage of S. Typhi.

CONCLUSIONS: Since 2011, there has been a rapid increase in the incidence of multidrug resistant, H58-lineage Typhoid in Blantyre. This is one of a number of reports of the re-emergence of Typhoid in Southern and Eastern Africa. There is an urgent need to understand the reservoirs and transmission of disease and how to arrest this regional increase.

Feasibility assessment of sugarcane expansion in southwest Goiás, Brazil based on the GIS technology.

The current study approaches the sugarcane culture expansion in Southwestern Goiás, especially in Mineiros, Quirinópolis and Rio Verde counties, which represent different times and responses to this process. The current logistics structure and future prospects for sugarcane and its derivatives transportation are studied at national level with emphasis to the aforementioned micro-region. Maps showing land use and land cover in three different years were generated from Landsat TM-5 satellite images and they were used to analyze the dynamics of changes in land use and in land cover. The region is marked by strong and rapid growth in the agricultural sector and its sugar-energy industry has been expanding in recent years, although with different aspects among its counties. Since it is a promising region in this sector, due to the favorable soil and weather conditions to the crop, the region requires more investment and planning in logistics to ensure production flow and to make it stronger within domestic and foreign markets.