In situ forming biodegradable hydrogels and their application for protein delivery

Hydrogels have been widely applied for biomedical applications, such as protein delivery and tissue engineering, due to their similarity with the extracellular matrix. Hydrogels are water-swollen, insoluble polymer networks. Their high water content renders them compatible with living tissue and proteins and their rubbery nature minimizes damage to the surrounding tissue. Conventionally, hydrogels are preformed and implanted in the body. More recently, hydrogels have been formed in situ under physiological conditions by mixing liquid precursors. These hydrogels are preferred over preformed hydrogels, since cells and bioactive compounds may be easily mixed with the precursor\ud solutions prior to gelation. Moreover, in situ gelation allows for minimally invasive surgery and for the preparation of complex shapes. Hydrogels are formed by physical or chemical crosslinking. Physical crosslinks are\ud formed by noncovalent interactions, such as hydrophobic and ionic interactions and stereocomplexation. Physical crosslinking generally proceeds under mild conditions, thus enabling in situ hydrogel formation and allowing the entrapment of labile compounds. The integrity of physically crosslinked hydrogels may however be lost upon a change in physical conditions. Chemically crosslinked hydrogels are formed by covalent bonds by reaction between functional groups. Most commonly, these hydrogels have been formed by radical chain polymerization of (meth)acrylate derived polymers initiated by photoirradiation. Chemically crosslinked hydrogels are generally more stable than physically crosslinked hydrogels. Chemically crosslinked hydrogels may also be formed in situ. However, care has to be taken that the reactants, products and/or auxiliary compounds are non-toxic

Efficacy of smoking prevention program 'Smoke-free Kids': study protocol of a randomized controlled trial

Contains fulltext : 77005.pdf (publisher's version ) (Open Access)Background - A strong increase in smoking is noted especially among adolescents. In the Netherlands, about 5% of all 10-year olds, 25% of all 13-year olds and 62% of all 17-year olds report ever smoking. In the U.S., an intervention program called 'Smoke-free Kids' was developed to prevent children from smoking. The present study aims to assess the effects of this home-based smoking prevention program in the Netherlands. Methods - A randomized controlled trial is conducted among 9 to 11-year old children of primary schools. Participants are randomly assigned to the intervention and control conditions. The intervention program consists of five printed activity modules designed to improve parenting skills specific to smoking prevention and parent-child communication regarding smoking. These modules will include additional sheets with communication tips. The modules for the control condition will include solely information on smoking and tobacco use. Initiation of cigarette smoking (first instance of puffing on a lighted cigarette), susceptibility to cigarette smoking, smoking-related cognitions, and anti-smoking socialization will be the outcome measures. To collect the data, telephone interviews with mothers as well as with their child will be conducted at baseline. Only the children will be examined at post-intervention follow-ups (6, 12, 24, and 36 months after the baseline). Discussion - This study protocol describes the design of a randomized controlled trial that will evaluate the effectiveness of a home-based smoking prevention program. We expect that a significantly lower number of children will start smoking in the intervention condition compared to control condition as a direct result of this intervention. If the program is effective, it is applicable in daily live, which will facilitate implementation of the prevention protocol. Trial registration: Netherlands Trial Register NTR146510 p

Enzyme-mediated fast in situ formation of hydrogels from dextran–tyramine conjugates

Dextran hydrogels were formed in situ by enzymatic crosslinking of dextran-tyramine conjugates and their mechanical, swelling and degradation properties were evaluated. Two types of dextran–tyramine conjugates (Mn,dextran=14 k, Mw/Mn=1.45), i.e. dextran–tyramine linked by a urethane bond (denoted as Dex–TA) or by an ester-containing diglycolic group (denoted as Dex–DG–TA), with different degrees of substitution (DS) were prepared. Hydrogels were rapidly formed under physiological conditions from Dex–TA DS 10 or 15 and Dex–DG–TA DS 10 at or above a concentration of 2.5 wt% in the presence of H2O2 and horseradish peroxidase (HRP). The gelation time ranged from 5 s to 9 min depending on the polymer concentration and HRP/TA and H2O2/TA ratios. Rheological analysis showed that these hydrogels are highly elastic. The storage modulus (G′), which varied from 3 to 41 kPa, increased with increasing polymer concentration, increasing HRP/TA ratio and decreasing H2O2/TA ratio. The swelling/degradation studies showed that under physiological conditions, Dex–TA hydrogels are rather stable with less than 25% loss of gel weight in 5 months, whereas Dex–DG–TA hydrogels are completely degraded within 4–10 d. These results demonstrate that enzymatic crosslinking is an efficient way to obtain fast in situ formation of hydrogels. These dextran-based hydrogels are promising for use as injectable systems for biomedical applications including tissue engineering and protein delivery

Stereocomplex Mediated Gelation of PEG-(PLA)2 and PEG-(PLA)8 Block Copolymers

Stereocomplex mediated hydrogels have been prepared by mixing solutions of polymers of opposite chirality of either PEG-(PLA)2 triblock copolymers or PEG-(PLA)8 star block copolymers. The critical gel concentrations of the mixed enantiomer solutions were considerably lower compared to polymer solutions containing only the single enantiomer. Moreover, gel-sol transition temperatures were increased and gel regions were expanded due to stereocomplexation. Rheology measurements showed that stereocomplexed hydrogels based on PEG-(PLA)8 have higher storage moduli compared to those based on PEG-(PLA)2. Stereocomplexed hydrogels prepared from 13 wt% PEG-(PLA)2 solutions in PBS showed a storage modulus of 0.9 kPa at 37 °C, while at similar conditions stereocomplexed hydrogels of PEG-(PLA)8 showed a storage modulus of 1.9 kPa at 10 wt%

Rapidly in Situ Forming Biodegradable Robust Hydrogels by Combining Stereocomplexation and Photopolymerization

Our previous studies have shown that stereocomplexed hydrogels can be rapidly formed in vitro as well as in vivo upon mixing aqueous solutions of eight-arm poly(ethylene glycol)−poly(l-lactide) (PEG−PLLA) and poly(ethylene glycol)−poly(d-lactide) (PEG−PDLA) star block copolymers. In this study, stereocomplexation and photopolymerization are combined to yield rapidly in situ forming robust hydrogels. Two types of methacrylate-functionalized PEG−PLLA and PEG−PDLA star block copolymers, PEG−PLLA−MA and PEG−PDLA−MA, which have methacrylate groups at the PLA chain ends and PEG−MA/PLLA and PEG−MA/PDLA, which have methacrylate groups at the PEG chain ends, were designed and prepared. Results showed that stereocomplexed hydrogels could be rapidly formed (within 1−2 min) in a polymer concentration range of 12.5−17.5% (w/v), in which the methacrylate group hardly interfered with the stereocomplexation. When subsequently photopolymerized, these hydrogels showed largely increased storage moduli as compared to the corresponding hydrogels that were cross-linked by stereocomplexation or photopolymerization only. Interestingly, the storage modulus of stereocomplexed−photopolymerized PEG−PLA−MA hydrogels increased linearly with increasing stereocomplexation equilibration time prior to photopolymerization (from ca. 6 to 32 kPa), indicating that stereocomplexation aids in photopolymerization. Importantly, photopolymerization of stereocomplexed hydrogels could take place at very low initiator concentrations (0.003 wt %). Swelling/degradation studies showed that combining stereocomplexation and photopolymerization yielded hydrogels with prolonged degradation times as compared to corresponding hydrogels cross-linked by photopolymerization only (3 vs 1.5 weeks). Stereocomplexed−photopolymerized PEG−MA/PLA hydrogels degraded much slower than corresponding PEG−PLA−MA hydrogels, with degradation times ranging from 7 to more than 16 weeks. Therefore, combining stereocomplexation and photopolymerization is a novel approach to obtain rapidly in situ forming robust hydrogels

Release of model proteins and basic fibroblast growth factor from in situ forming degradable dextran hydrogels

Our previous studies showed that degradable dextran hydrogels are rapidly formed in situ upon mixing aqueous solutions of dextran vinyl sulfone (dex-VS) conjugates and tetrafunctional mercapto poly(ethylene glycol) (PEG-4-SH) by Michael addition. The hydrogel degradation time and storage modulus could be controlled by the degree of vinyl sulfone substitution (DS) and dextran molecular weight. The degradation time could further be adjusted by the spacer between the thioether and the ester bond of the dex-VS conjugates (ethyl vs. propyl, denoted as dex-Et-VS and dex-Pr-VS, respectively). In this paper, the release of three model proteins, i.e. immunoglobulin G (dh = 10.7 nm, IgG), bovine serum albumin (BSA, dh = 7.2 nm) and lysozyme (dh = 4.1 nm), as well as basic fibroblast growth factor (bFGF) from these in situ forming dextran hydrogels is studied. Proteins could be easily loaded into the hydrogels by mixing protein containing solutions of dex-VS and PEG-4-SH. The release of IgG from dex-Et-VS hydrogels followed biphasic release kinetics, with a slow, close to first order release for the first 9 days followed by an accelerated release and over 80% of IgG was released in 12 to 25 days. Interestingly, the release of IgG from dex-Pr-VS hydrogels followed close to zero order kinetics, wherein approximately 95% was released in 21 days. The release of BSA from dex-Pr-VS hydrogels followed biphasic kinetics, with almost first order release followed by close to zero order release. Approximately 75% of the entrapped BSA could be released from dex-Pr-VS hydrogels in 16 days. Dex-Pr-VS hydrogels released 40% of lysozyme in 14 days, with full preservation of the enzymatic activity of the released lysozyme, as determined by bacteria lysis experiments. The release of basic fibroblast growth factor (bFGF) from dex-Pr-VS hydrogels showed first order kinetics, with quantitative release in 28 days. These results show that the in situ forming degradable dextran hydrogels can be used for the controlled release of proteins

Novel in Situ Forming, Degradable Dextran Hydrogels by Michael Addition Chemistry: Synthesis, Rheology, and Degradation

Various vinyl sulfone functionalized dextrans (dex-VS) (Mn,dextran = 14K or 31K) with degrees of substitution (DS) ranging from 2 to 22 were conveniently prepared by a one-pot synthesis procedure at room temperature. This procedure involved reaction of a mercaptoalkanoic acid with an excess amount of divinyl sulfone yielding vinyl sulfone alkanoic acid, followed by conjugation to dextran using N,N‘-dicyclohexylcarbodiimide (DCC)/4-(dimethylamino)pyridinium 4-toluenesulfonate (DPTS) as a catalyst system. By using two different mercaptoalkanoic acids, 3-mercaptopropionic acid (1a) and 4-mercaptobutyric acid (1b), dex-VS conjugates with either an ethyl spacer (denoted as dex-Et-VS) or a propyl spacer (denoted as dex-Pr-VS) between the thioether and ester groups were obtained. Linear and four-arm mercaptopoly(ethylene glycol) (Mn = 2.1K) with two or four thiol groups (denoted as PEG-2-SH and PEG-4-SH, respectively) were also prepared. Hydrogels were rapidly formed in situ under physiological conditions by Michael type addition upon mixing aqueous solutions of dex-VS and multifunctional PEG-SH at a concentration of 10−20% w/v. The gelation time ranged from 0.5 to 7.5 min, depending on the DS, concentration, dextran molecular weight, and PEG-SH functionality. Rheological studies showed that these dextran hydrogels are highly elastic. The storage modulus increased with increasing DS, concentration, and dextran molecular weight, and hydrogels with a broad range of storage moduli from 3 to 46 kPa were obtained. Swelling/degradation studies revealed that these dextran hydrogels have a low initial swelling and are degradable under physiological conditions. The degradation time varied from 3 to 21 days depending on the DS, concentration, dextran molecular weight, and PEG-SH functionality. Interestingly, dex-Pr-VS hydrogels showed prolonged degradation times, but otherwise similar properties compared to dex-Et-VS hydrogels. The hydrolysis of the linker ester bonds of the dex-VS conjugates under physiological conditions was confirmed by 1H NMR. The results showed that the hydrolysis kinetics were independent of the DS and the dextran molecular weight. Therefore, the degradation rate of these hydrogels can be precisely controlled