Reference gene selection in human periodontal ligament cells

Background: Real-time quantitative PCR (qPCR) is a highly sensitive and specific method which is used extensively for determining gene expression profiles in a variety of cell and tissue types. In order to obtain accurate and reliable gene expression quantification, qPCR data are generally normalised against so-called reference or housekeeping genes. Ideally, reference genes should have abundant and stable RNA transcriptomes under the experimental conditions employed. However, reference genes are often selected rather arbitrarily and indeed some have been shown to have variable expression in a variety of in vitro experimental conditions.
Objective: The objective of the current study was to investigate reference gene expression in human periodontal ligament (PDL) cells in response to treatment with lipopolysaccharide (LPS).
Method: Primary human PDL cells were grown in Dulbecco’s Modified Eagle Medium with L-glutamine supplemented with 10% fetal bovine serum, 100UI/ml penicillin and 100µg/ml streptomycin. RNA was isolated using the RNeasy Mini Kit (Qiagen) and reverse transcribed using the QuantiTect Reverse Transcription Kit (Qiagen). The expression of a total of 19 reference genes was studied in the presence and absence of LPS treatment using the Roche Reference Gene Panel. Data were analysed using NormFinder and Bestkeeper validation programs.
Results: Treatment of human PDL cells with LPS resulted in changes in expression of several commonly used reference genes, including GAPDH. On the other hand the reference genes β-actin, G6PDH and 18S were identified as stable genes following LPS treatment.
Conclusion: Many of the reference genes studied were robust to LPS treatment (up to 100 ng/ml). However several commonly employed reference genes, including GAPDH varied with LPS treatment, suggesting they would not be ideal candidates for normalisation in qPCR gene expression studies.

Gingival fibroblasts respond to the TRPV1 agonist capsaicin

Background: The oral cavity is a frontline barrier which is often exposed to physical trauma and noxious substances, leading to pro-inflammatory responses designed to be protective in nature. The transient receptor potential (TRP) super family of ion channels is believed to play a critical role in sensory physiology, acting as transducers for thermal, mechanical and chemical stimuli. Our understanding of the role of TRP channel activation in gingival and periodontal inflammation is currently limited. Gingival fibroblasts are the most abundant structural cell in periodontal tissues and we hypothesised that they may have a role in the inflammatory response associated with TRP channel activation. Objectives: The present study was designed to determine whether the TRPV1 agonist capsaicin could elicit a pro-inflammatory response in gingival fibroblasts in vitro by up-regulation of interleukin-8 (IL-8) production. Methods: Gingival fibroblasts were derived by explant culture from surgical tissues following ethical approval. Cells were maintained in Dulbecco's modified Eagle's medium (DMEM), containing 10% fetal calf serum (FCS) in 5% CO2. Following treatment of gingival fibroblasts with capsaicin, IL-8 levels were measured by ELISA. The potential cytotoxicity of capsaicin was determined by the MTT assay. Results: In gingival fibroblasts treated with the TRPV1 agonist capsaicin (10µM), IL-8 production was significantly increased compared with untreated control cells. Capsaicin was shown not to be toxic to gingival fibroblasts at the concentrations studied. Conclusion: The identification of factors that modulate pro-inflammatory cytokine production is important for our understanding of gingival and periodontal inflammation. This study reports for the first time that gingival fibroblasts respond to the TRPV1 agonist capsaicin by increased production of IL-8. Activation of TRPV1 on gingival fibroblasts could therefore have an important role in initiating and sustaining the inflammatory response associated with periodontal diseases

Functional expression of TRPV4 in human periodontal ligament cells

Background: Periodontal ligament (PDL) cells are exposed to physical forces in vivo in response to mastication, parafunction, speech and orthodontic tooth movement. Although it has been shown that PDL cells perceive and respond directly to mechanical stimulation, the nature of the ion channels that mediate this mechanotransduction remain to be fully elucidated. The transient receptor potential (TRP) superfamily of ion channels is believed to play a critical role in sensory physiology, where they act as transducers for thermal, chemical and mechanical stimuli. Recent studies have shown that members of the vanilloid (TRPV) and ankyrin (TRPA) subfamilies encode mechanosensitive TRPs. The vanilloid family member TRPV4 is one such non selective calcium permeable cationic channel which has been shown to be activated by chemical ligands, hypotonicity, and mechanical stimuli. Objectives: The objective of the current study was to investigate functional expression of TRPV4 in cultured human PDL cells. Methods: Human PDL cells were grown in Dulbecco's Modified Eagle Medium with L-glutamine supplemented with 10% fetal bovine serum (FBS), 100UI/ml penicillin and 100μg/ml streptomycin. Cells in passage 4-6 were used in all experiments. TRPV4 functional expression was determined using ratiometric calcium imaging. Cultured cells were loaded with intracellular Ca2+ probe fura-2 and cells were then stimulated with the TRPV4 agonists, 4alpha-phorbol 12,13-didecanoate (4alpha-PDD), GSK1016790A or hypotonic solution. The TRPV4 antagonist RN 1734 was used to block the corresponding agonist responses. Results: PDL fibroblasts responded to application of TRPV4 agonists and hypotonic stimuli by an increase in intracellular calcium which was attenuated in the presence of the TRPV4 antagonist. Conclusions: We have shown for the first time the functional expression of the mechanosensitive TRPV4 channel in human PDL cells. The molecular identity and mechanisms of activation of mechanosensitive TRP channels in PDL cells merit further investigation.

Vom Trümmerfilm zur Traumfabrik : die "Junge Film-Union". Eine Fallstudie zur westdeutschen Filmproduktion

Die Trümmer des deutschen Reiches waren 1945 auch die Trümmer des deutschen Films. Der Neuaufbau der deutschen Filmwirtschaft, der unter der Ägide der Alliierten begann, eröffnete trotz zahlloser Schwierigkeiten neue Chancen, nicht zuletzt regional: auch Niedersachsen wurde "Filmland". Die "Junge Film-Union", 1947 lizensiert, wurde bald zur größten niedersächsischen Filmproduktionsfirma. Waren die ersten zeitnahen Filme - auch "Trümmerfilme" genannt - noch in mancher Hinsicht improvisiert, so professionalisierte sich die Produktion zusehends. Es wurden eigene Ateliers gebaut, in dem Heideort Bendestorf entstand in kurzer Zeit eine Filmindustrie. Und auch die Filmstoffe wandelten sich, ebenso die Besetzung der Hauptrollen: der "zeitlose" Unterhaltungsfilm wurde selbstverständlich, ein neuer Kult mit alten Stars verdrängte den Nachwuchs. Neben vielen Mißerfolgen der "Jungen Film-Union" wurde schließlich "Die Sünderin" zu einem (Skandal-)Erfolg, der Marika-Rökk-Film "Die Csardas-Fürstin" ein echter Publikumsrenner. Der spektakuläre Neuaufbau dieser Filmproduktion, die wechselvolle Entwicklung, die im Konkurs 1952 ein jähes Ende fand, werfen Fragen auf, denen in dieser Arbeit auf der Basis neuen Quellenmaterials nachgegangen wird. Welche politischen, ökonomischen und kulturellen Einflüsse waren es, die für die "Junge Film-Union" bestimmend wurden. Welche Rolle spielte die britische Filmpolitik? Welche Bedeutung hatte die Währungsreform, wie wirkten sich die Bundesbürgschaften aus? Welche filmischen Traditionen wurden aufgegriffen, und wie spiegelt sich die außerfilmische Realität in den Produktionen? Die Ergebnisse der Untersuchung werfen dabei auch ein Licht auf den Neuaufbau der westdeutschen Filmwirtschaft insgesamt.

Inspecção-Geral de Educação

Relatório de estágio de mestrado, Ciências da Educação (Administração Educacional), Universidade de Lisboa, Instituto de Educação, 2011

Circulation patterns and processes in Barkley Sound, Vancouver, BC

Senior thesis written for Oceanography 444

Survey of dinoflagellate cysts in Barkley Sound, Vancouver Island, Canada

Senior thesis written for Oceanography 445

Ecologia alimentar e biometria de duas espécies de raias de espinho dos Açores : uma análise ecológica e perspetiva para turismo subaquático

Dissertação de Mestrado em Gestão e Conservação da Natureza.

Brock University Coat of Arms

The armorial bearings for Brock University, or more simply the University Coat of Arms, or crest, was designed in a large part by Presdent Gibson, assisted by other members of the Board of Governors (A preliminary design can be seen here). The Coat of Arms was granted to Brock University on March 17th, 1965. The Coat of Arms consist of an eagle, taken from General Brock’s own arms, displayed against a scarlet background - one of the official colours of Brock University. Immediately above it on a chief argent is displayed a maple-leaf (for Canada), a scallop shell (from the Lincoln and Welland regiment), and a trillium (for the province of Ontario). An open book fronts the eagle representing learning and knowledge. The crest itself is made up of a torch symbolizing learning, surrounded by a serpent for wisdom, with two calumet or North American pipes of peace, to symbolize Canada, friendship and agreement. The supporters consist of a beaver on the dexter side, emblematic of Canada and representing work and industry in learning. On the sinister side, a brock or badger (also in commemoration of General Brock) represents tenacity of purpose. The motto 'Surgite' is visible just below the arms.

Office of the City Clerk, St. Catharines Records, 1928-1974, n.d.

The site of present-day St. Catharines was settled by 3000 United Empire Loyalists at the end of the 18th century. From 1790, the settlement (then known as "The Twelve") grew as an agricultural community. St. Catharines was once referred to Shipman's Corners after Paul Shipman, owner of a tavern that was an important stagecoach transfer point. In 1815, leading businessman William Hamilton Merritt abandoned his wharf at Queenston and set up another at Shipman's Corners. He became involved in the construction and operation of several lumber and gristmills along Twelve Mile Creek. Shipman's Corners soon became the principal milling site of the eastern Niagara Peninsula. At about the same time, Merritt began to develop the salt springs that were discovered along the river which subsequently gave the village a reputation as a health resort. By this time St. Catharines was the official name of the village; the origin of the name remains obscure, but is thought to be named after Catharine Askin Robertson Hamilton, wife of the Hon. Robert Hamilton, a prominent businessman. Merritt devised a canal scheme from Lake Erie to Lake Ontario that would provide a more reliable water supply for the mills while at the same time function as a canal. He formed the Welland Canal Company, and construction took place from 1824 to 1829. The canal and the mills made St. Catharines the most important industrial centre in Niagara. By 1845, St. Catharines was incorporated as a town, with the town limits extending in 1854. Administrative and political functions were added to St. Catharines in 1862 when it became the county seat of Lincoln. In 1871, construction began on the third Welland Canal, which attracted additional population to the town. As a consequence of continual growth, the town limits were again extended. St. Catharines attained city status in 1876 with its larger population and area. Manufacturing became increasingly important in St. Catharines in the early 1900s with the abundance of hydro-electric power, and its location on important land and water routes. The large increase in population after the 1900s was mainly due to the continued industrialization and urbanization of the northern part of the city and the related expansion of business activity. The fourth Welland Canal was opened in 1932 as the third canal could no longer accommodate the larger ships. The post war years and the automobile brought great change to the urban form of St. Catharines. St. Catharines began to spread its boundaries in all directions with land being added five times during the 1950s. The Town of Merritton, Village of Port Dalhousie and Grantham Township were all incorporated as part of St. Catharines in 1961. In 1970 the Province of Ontario implemented a regional approach to deal with such issues as planning, pollution, transportation and services. As a result, Louth Township on the west side of the city was amalgamated, extending the city's boundary to Fifteen Mile Creek. With its current population of 131,989, St. Catharines has become the dominant centre of the Niagara region. Source: City of St. Catharines website http://www.stcatharines.ca/en/governin/HistoryOfTheCity.asp (January 27, 2011)

Second Welland Canal - Book 3, Survey Map 13 - Village of Welland - Merrittville and Crowland

Survey map of the Second Welland Canal created by the Welland Canal Company showing the township of Crowland as well as the Village of Welland-Merritville. Identified structures associated with the Canal include Old Canal, old towing path, water way, bridge, culvert, covered drain, drain, and towing path. Surveyor measurements and notes can be seen in red and black ink and pencil. Local area landmarks include E. Seeley's store, W.A. Bald's store, barn, and two ponds. Roads parallel to Canal include the old towing road, towing path, Canal Street, road to Welland_lle, and road to Junction. Roads perpendicular to Canal include Road Allowance between the 5th and 6th Concession, Division Street, Road to Narrows. Properties and property owners are noted as W.A. Bald, E. Seeley, John Price, Eli Mead, Jacob Griffith, John Hellems. Lots noted are: Lots Number 25, 26, 5th Concession.

The Columbian Newspaper vol. IV no. 1105- June 12, 1813

The "Columbian" was established by Mr. Holt in 1808 as an "organ of the Clintonians" (Journalism in the United States, from 1690-1872 By Frederic Hudson) and a replacement to the "Citizen". It was a passionate Democratic paper that supported both Jefferson and Madison. Holt later sold the "Columbian" to Alden Spooner, who published the Long Island "Star". The Columbian ceased in 1821. This issue includes the following War of 1812 topics: Page 2: copies of letters from U.S. Gen. H. Dearborn describing the U.S. attack on Ft. George 27 May 1813; letter from Gen. H. Dearborn describing U.S. pursuit of British troops at Beaver Dams and the U.S. capture of Fort Erie; list of U.S. killed and wounded in Commodore Isaac Chauncey's squadron in attack on York 27 April, 1813; list of killed and wounded in Commodore Isaac Chauncey's squadron in attack on Ft. George 27 May 1813; abstract of the cartel for the exchange of prisoners of war between Great Britain and the United States; mention of troop movements through New York on their way to the battle front; list of persons killed or lost on the privateer Saratoga; Page 3: account of the loss of the U.S. war ships the Growler and Eagle to the British; news from Quebec and Kingston of troop movements; U.S. ship Syren unsuccessfully pursues British ship Herald; account of a U.S. cargo ship being captured by a British military ship; account of U.S. ship Siro capturing British ship Loyal Sam; report from Halifax of recent British troop and ship movements; list of recent troop enrollments in various states; report of British troop build up in Kingston;

Personal Seals of Sir Isaac Brock

Two orange coloured seals containing the impression of the personal seal of Sir Isaac Brock. The seal depicts a crest containing a lion in the upper portion and an eagle with its wings spread in the lower portion, which represent strength, clarity of vision and courage. Below the crest is a banner with "Canada" written on it. Above the crest is an Indian on a battlement holding a tomahawk, which represents the alliance Brock had forged with the First Nations. The seals are in a John Sinclair tobacco tin.

The Washingtonian, 17 May 1813 (vol. 3, no. 148) - Newspaper

Volumes of interest were published between 1812 and 1815 with articles about the War of 1812. Issue for July 12, 1813 includes an account of the capture of the Chesapeake, report of explosion of the schooner Eagle off New London, report Com. Hardy had detained a smack, was determined to destroy all smacks and other small craft he found, reports of recent events at Malden, Fort George, head of the Lake, Sacketts Harbour, two reports of attack on Hampton, report of British attack near point of Nansemond River/Craney Island, second report of battle on/near Craney Island, additional report of attack at Hampton, marine news from Baltimore, report that sickness rampant throughout Army, many deaths reported, report that British were fortifying Brownstown, but deserters from British believe Michigan could be retaken by 3000 troops, report Gen. Boyd remained at Fort George to supervise repair of fortifications, report British reinforced from Kingston in Niagara and in Malden, report Gen. Hampton arrived at Albany on way to Burlington, report Gen. Parker left Albany, headed to Burlington, report of 400 men marching from Greenbush to the frontier, report of battles at Stoney Creek, 40 Mile Creek, report that Gen. Boyd lost an action with British in which 8 to 10 hundred men killed and taken from Americans, reports British had captured stores at Black Rock, Sodus, and Oswego, report of British victory in a battle 5 miles from Queenston, troop strength under Dearborn in Niagara reported between 2 and 3 thousand, additional reports of Battle of Beaverdams, reports Dearborn had resumed command, after lengthy illness, Gen. Boyd remained at Fort George, reports natives had been sieging Fort Wayne.

The Columbian, 12 June 1813 (vol. 4, no. 1105) - Newspaper

Volumes of interest were published between 1812 and 1815 with articles about the War of 1812. Issue for June 12, 1813 includes copies of letters from U.S. Gen. H. Dearborn describing the U.S. attack on Ft. George 27 May 1813; letter from Gen. H. Dearborn describing U.S. pursuit of British troops at Beaver Dams and the U.S. capture of Fort Erie; list of U.S. killed and wounded in Commodore Isaac Chauncey’s squadron in attack on York 27 April, 1813; list of killed and wounded in Commodore Isaac Chauncey’s squadron in attack on Ft. George 27 May 1813; abstract of the cartel for the exchange of prisoners of war between Great Britain and the United States; mention of troop movements through New York on their way to the battle front; list of persons killed or lost on the privateer Saratoga; account of the loss of the U.S. war ships the Growler and Eagle to the British; news from Quebec and Kingston of troop movements; report of launching of British ship Sir George Prevost; announcement of sending of 5 American infantry companies to Burlington from Bennington, Vermont; U.S. ship Syren unsuccessfully pursues British ship Herald; account of a U.S. cargo ship being captured by a British military ship; account of U.S. ship Siro capturing British ship Loyal Sam; report from Halifax of recent British troop and ship movements; list of recent troop enrollments in various states; report of British troop build up in Kingston.