Table 2.

Researches on natural polymer-based layered scaffolds for periodontal regeneration.

Biomaterials	Target periodontal tissue	Bilayered/Trilayered/method of fabrication	Significant results	Ref
Non-cross linked collagen type I and III	Soft tissue (gingiva, PDL) Hard tissue (alveolar bone, cementum)	Bilayered lyophilization	Combination of collagen I and III: improve the stability, mechanical properties, and cellular properties No chemical crosslinking: improve the cell attachment and proliferation Low porosity, smooth and thin layer with elastic properties improves the suturing of host mucosal margins High porosity layer increases the tissue adherence, improve cell integration, and improve wound healing process. High porosity side can face to bone or soft tissue and improve each side regeneration	68
Non-cross linked collagen type I and III	Soft tissue (gingiva PDL) Hard tissue (alveolar bone, cementum)	Bilayered Lyophilization	One dense layer and high porosity layer to improve cell attachment Use two different positions, one dense layer faces with soft tissue and other one upside-down same Radiological and histomorphometric results in both positions show no orientation preference in bone defects	69
Collagen and calcium silicate with strontium doped	Hard tissue (Alveolar bone and cementum)	Bilayered 3D printing	Calcium silicate (CS): increases the bonding between surrounding bone and new scaffolds because of hydroxyapatite formation on the surface of scaffold, promoting the dentin metabolism and increasing secretion of cementum, supporting bone tissue for soft tissue was formed great bone formation of bilayered cell laden structure after 12 weeks Excellent improvement in bone formation in presence of Sr significant improvement in cell laden bilayered scaffold (∼20%) while bone volume fraction in nest bilayered scaffold is 13% and in SrCS is 9%. Higher and trabecular thickness in cell laden bilayered scaffold is comparable to others	70
Different molecular wight Chitosan with genipin crosslinking	Alveolar bone, gingiva and PDL	trilayered freeze drying	Different molecular weight chitosan: match degradation rate and mechanical properties with target tissue Controllable degradation rate and great PDL regeneration	71
Chitosan membranes with Doxycycline hyclate	Soft tissue (gingiva, PDL) Hard tissue (alveolar bone, cementum)	Bilayered and trilayered	Doxycycline hyclate: decrease the bacteria infection in the periodontal defect site Suitable drug release especially in the first stage and efficient dosage at long term Appropriate mechanical properties were seen	72
Collagen and chitosan First layer: two solid layers of chitosan. And collagen Second layer: electrospined collagen nanofiber on the chitosan sublayer	Hard tissue (alveolar bone, cementum)	Bilayered Electrospinning	Collagen: great biocompatibility, low tissue morbidity, good resorbability, bio-affinity, poor effective shield in bone defect, rapid degradation, early collapse, without any effective blood clot transformation into the bone significant increase in rabbit MSCs activity for 2 weeks More metabolic activity of MSCs cells after 3 days Higher cellular activities on the second layer due to higher surface area of collagen fibers considerable difference for Colα1 and Runx-2 between two layers after three weeks More bone formation and no inflammation responses were seen	73
Chitosan and gelatin	Soft tissue (gingiva, PDL)	Bilayered Solvent casting and freeze drying And chemical reaction between layers with genipin	Genipin: increasing the interactions between layer and increasing mechanical properties and stability Gelatin: increasing mechanical properties of chitosan membrane Suitable mechanical properties: Yield stress in the range of 10 kPa and 19 kPa, elastic modules: 26–34 kPa Rapid mineralization	74